{"id":31,"date":"2023-03-29T19:42:51","date_gmt":"2023-03-29T19:42:51","guid":{"rendered":"https:\/\/test-hcc-press-wp-multisite.pantheonsite.io\/bsc2010l\/?post_type=chapter&#038;p=31"},"modified":"2025-10-29T16:34:15","modified_gmt":"2025-10-29T16:34:15","slug":"metric-measurement-measuring-very-small-volume","status":"publish","type":"chapter","link":"https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/chapter\/metric-measurement-measuring-very-small-volume\/","title":{"raw":"Chapter 3 - Volume Measurements using Micropipettes","rendered":"Chapter 3 &#8211; Volume Measurements using Micropipettes"},"content":{"raw":"<h2>Volume Measurements using Micropipettes<\/h2>\r\n<h3>BACKGROUND<\/h3>\r\nVery tiny amounts of chemicals and biological reagents are used in many molecular biology experiments. To measure these minute [pb_glossary id=\"334\"]volumes[\/pb_glossary], technicians use [pb_glossary id=\"528\"]micropipettes[\/pb_glossary] that measure in [pb_glossary id=\"348\"]microliter[\/pb_glossary] (\u00b5l) amounts. This [pb_glossary id=\"515\"]metrology[\/pb_glossary] activity introduces micropipetting technique. As with all fine motor skills, learning how to use a micropipette takes practice and determination. Students must be able to measure these very tiny volumes with [pb_glossary id=\"520\"]accuracy[\/pb_glossary] and [pb_glossary id=\"524\"]precision[\/pb_glossary].\r\n\r\nMicropipettes are used to dispense solutions in the [pb_glossary id=\"348\"]microliter[\/pb_glossary] range. The microliter is a small volume, yet measurable in a typical laboratory. A [pb_glossary id=\"534\"]milliliter[\/pb_glossary] is a metric unit of volume equal to a thousandth of a liter. A microliter is a unit of volume in the Metric System. The symbol for microliter is \u03bcL. There are 1,000 microliters in a [pb_glossary id=\"534\"]milliliter[\/pb_glossary]. An example of when you might use microliter volumes would be in preparation of an electrophoresis sample, when isolating DNA, or during chemical purification.\r\n\r\nVolume smaller than microliter is [pb_glossary id=\"352\"]nanoliter[\/pb_glossary]. The prefix nano means one billionth and is symbolized as n in the [pb_glossary id=\"537\"]Metric System[\/pb_glossary]. Nano denotes a factor of one billionth (10<sup>-9<\/sup>) which means that there are 1,000,000,000 nanoliters in a [pb_glossary id=\"517\"] liter[\/pb_glossary].\r\n\r\nUse the following information to calculate metric volume conversions:\r\n<ul>\r\n \t<li>1 \u00b5L = 10<sup>\u20136<\/sup> L = 10<sup>\u20133<\/sup> mL<\/li>\r\n \t<li>1 nL = 10<sup>\u20139<\/sup><span style=\"font-size: 1em;\"> L = 10<\/span><sup>\u20136<\/sup><span style=\"font-size: 1em;\"> mL<\/span><\/li>\r\n \t<li>1000 nL = 1 \u00b5L<\/li>\r\n \t<li>0.001 \u00b5L = 1 nL<\/li>\r\n<\/ul>\r\n[pb_glossary id=\"528\"]Micropipettes[\/pb_glossary] are among the<span class=\"ls0\">\u00a0essential laboratory instrument used <\/span>to accurately distribute different volumes of liquids in the microliter range. \u00a0In order to prevent cross contamination<span class=\"ls3 ws1\">, m<span class=\"ls0\">icropipettes use disposable tips to aspirate liquid and distribute<\/span><\/span> the solution.Micropipettes enable sterile and accurate liquid handling.\r\n\r\nThere are several\u00a0types of micropipettes\u00a0on the market which can be classified according to:\r\n<ul>\r\n \t<li>Volume: fixed or variable volume\u00a0micropipette<\/li>\r\n \t<li>The principle of operation: air displacement or positive displacement\u00a0micropipette.<\/li>\r\n \t<li>The operating mechanism: mechanical or electronic micropipette<\/li>\r\n \t<li>The number of channels: single-channel or multi-channel\u00a0micropipette.<\/li>\r\n<\/ul>\r\nThe different parts of a variable volume micropipette are:\r\n<ul>\r\n \t<li>Plunger that is used for volume adjustment and to aspirate and dispense the desired amount of liquid into the tip of the micropipette.<\/li>\r\n \t<li>Trip Ejector that helps to remove the tip from the micropipette, without touching them, by pressing the tip eject button.<\/li>\r\n \t<li>Volume Window that displays the adjusted volume in the volume window (volume set to be aspirated\/dispensed).<\/li>\r\n \t<li>Micropipette Shaft is a tube filled with air. Pressing the piston expels a volume of air contained in the rod, releasing the piston allows this air to return to the rod.<\/li>\r\n \t<li>Micropipette Tips are tips attached to a micropipette to collect liquid and then transfer it from one place to another. Tips of different sizes are used to collect different volumes of liquid.<\/li>\r\n<\/ul>\r\n[caption id=\"attachment_343\" align=\"aligncenter\" width=\"1024\"]<img class=\"wp-image-343 size-large\" src=\"http:\/\/pressbooks.hcfl.edu\/bsc2010l\/wp-content\/uploads\/sites\/22\/2023\/03\/P1000-1024x576.png\" alt=\"This is a P -1000 micropipette illustrating the different parts of the instrument. Long description is included in the text.\" width=\"1024\" height=\"576\" \/> P-1000 Micropipette[\/caption]\r\n\r\n[caption id=\"attachment_341\" align=\"aligncenter\" width=\"1024\"]<img class=\"wp-image-341 size-large\" src=\"http:\/\/pressbooks.hcfl.edu\/bsc2010l\/wp-content\/uploads\/sites\/22\/2023\/03\/P200-1024x576.png\" alt=\"This is a P- 200 micropipette illustrating the different parts of the instrument. Long description is included in the text.\" width=\"1024\" height=\"576\" \/> P-200 Micropipette[\/caption]\r\n\r\n[caption id=\"attachment_339\" align=\"aligncenter\" width=\"1024\"]<img class=\"wp-image-339 size-large\" src=\"http:\/\/pressbooks.hcfl.edu\/bsc2010l\/wp-content\/uploads\/sites\/22\/2023\/03\/P20-1024x576.png\" alt=\"This is a P-20 micropipette illustrating the different parts of the instrument. Long description is included in the text.\" width=\"1024\" height=\"576\" \/> P-20 Micropipette[\/caption]\r\n\r\nIn a variable volume micropipette the volume of liquid to be aspirated or dispensed are adjusted according to the capacity of the micropipette (a specific minimum and maximum volume range). The acronyms P10, P20, P1000 on micropipettes\u00a0represent the maximum volume that can be aspirated\/dispensed\r\n\r\nIn the lab, the most common micropipettes are:\r\n<ul>\r\n \t<li>P-10 measures volumes of 0.5 \u00b5l to 10 \u00b5l in 0.02 \u00b5l increments;<\/li>\r\n \t<li>P-20 measures volumes of 2 \u00b5l to 20 \u00b5l in 0.02 \u00b5l increments;<\/li>\r\n \t<li>P-200 measures volumes of 20 \u00b5l to 200 \u00b5l in 0.2 \u00b5l increments;<\/li>\r\n \t<li>P-1000 measures volumes of 100 \u00b5l to 1000 \u00b5l in 2 \u00b5l increments.<\/li>\r\n<\/ul>\r\n[caption id=\"attachment_493\" align=\"aligncenter\" width=\"400\"]<img class=\"wp-image-493\" src=\"http:\/\/pressbooks.hcfl.edu\/bsc2010l\/wp-content\/uploads\/sites\/22\/2023\/03\/Micopipette_calculation-300x158.png\" alt=\"This photo illustrates micropipetting volume measurements for P-20, P-200 and P-1000. More details are in the text.\" width=\"400\" height=\"210\" \/> Measuring Volumes with Different Micropipettes[\/caption]\r\n\r\nOperate the micropipette slowly and carefully.\r\n\r\nAlways use the correct pipet for the job. Most labs have four sizes of micropipettes:\r\n\r\nNever attempt to set a volume beyond the pipet\u2019s minimum or maximum values. Only set the volume within the range specified for that micropipette.\r\n<div class=\"row justify-content-around\">\r\n<div class=\"col-sm-7\">\r\n<table class=\"aligncenter\" style=\"height: 120px;\"><caption>Types of Micropipettes and Their Ranges<\/caption>\r\n<thead>\r\n<tr style=\"height: 15px;\">\r\n<th style=\"width: 163.479px; height: 15px; text-align: left;\" scope=\"col\">Acronym<\/th>\r\n<th style=\"width: 240.604px; height: 15px; text-align: left;\" scope=\"col\">Volume<\/th>\r\n<\/tr>\r\n<\/thead>\r\n<tbody class=\"text-center\">\r\n<tr style=\"height: 15px;\">\r\n<td style=\"width: 163.812px; height: 15px;\">P-2<\/td>\r\n<td style=\"width: 240.938px; height: 15px;\">0.2 \u2013 2 \u00b5l<\/td>\r\n<\/tr>\r\n<tr style=\"height: 15px;\">\r\n<td style=\"width: 163.812px; height: 15px;\">P-10<\/td>\r\n<td style=\"width: 240.938px; height: 15px;\">1 \u2013 10 \u00b5l<\/td>\r\n<\/tr>\r\n<tr style=\"height: 15px;\">\r\n<td style=\"width: 163.812px; height: 15px;\">P-20<\/td>\r\n<td style=\"width: 240.938px; height: 15px;\">2 \u2013 20 \u00b5l<\/td>\r\n<\/tr>\r\n<tr style=\"height: 15px;\">\r\n<td style=\"width: 163.812px; height: 15px;\">P-100<\/td>\r\n<td style=\"width: 240.938px; height: 15px;\">20 \u2013 100 \u00b5l<\/td>\r\n<\/tr>\r\n<tr style=\"height: 15px;\">\r\n<td style=\"width: 163.812px; height: 15px;\">P-200<\/td>\r\n<td style=\"width: 240.938px; height: 15px;\">20 \u2013 200 \u00b5l<\/td>\r\n<\/tr>\r\n<tr style=\"height: 15px;\">\r\n<td style=\"width: 163.812px; height: 15px;\">P-1000<\/td>\r\n<td style=\"width: 240.938px; height: 15px;\">100 \u2013 1000 \u00b5l<\/td>\r\n<\/tr>\r\n<\/tbody>\r\n<\/table>\r\n<\/div>\r\n<\/div>\r\n<span data-preserver-spaces=\"true\">Micropippete performance is evaluated using a\u00a0<\/span>[pb_glossary id=\"539\"]gravimetric method[\/pb_glossary]. <span data-preserver-spaces=\"true\">Gravimetric methods depend on the high [pb_glossary id=\"520\"]accuracy[\/pb_glossary] and [pb_glossary id=\"524\"]precision [\/pb_glossary]of analytical balances. During gravimetric testing, a desired volume of purified water is aspirated and dispensed from the device to be checked. The\u00a0<\/span><em><span data-preserver-spaces=\"true\">desired volume is termed the<\/span><\/em><span data-preserver-spaces=\"true\">\u00a0<\/span>\"nominal volume.<span data-preserver-spaces=\"true\">\" The volume [pb_glossary id=\"513\"]measurement[\/pb_glossary] is then checked by weighing the liquid on a high-quality analytical balance. A calculation is performed to convert the weight of the water to a volume. If the volume-measuring device is accurately operating, then the nominal volume will be identical to the volume calculated from the weight of the water. This process indicates the [pb_glossary id=\"520\"]accuracy[\/pb_glossary] of the device being tested.<\/span>\r\n\r\nOverview and demonstration of\u00a0 miciropipetting technique\r\n<ul>\r\n \t<li>Visit <a href=\"https:\/\/www.ncbionetwork.org\/educational-resources\/videos\/micropipetting\"><strong>NC BioNetwork Virtual Microscope<\/strong><\/a><\/li>\r\n \t<li>Visit <strong><a href=\"https:\/\/florida.pbslearningmedia.org\/resource\/biot11.sci.life.gen.usingmicro\/using-a-micropipette\/\">Florida PBS Learning Media<\/a><\/strong><\/li>\r\n<\/ul>\r\n<div class=\"textbox textbox--key-takeaways\"><header class=\"textbox__header\">\r\n<h4 class=\"textbox__title\">Key Terms<\/h4>\r\n<\/header>\r\n<div class=\"textbox__content\">\r\n<ul>\r\n \t<li>Metrology<\/li>\r\n \t<li>Measurements<\/li>\r\n \t<li>Liters, Milliliters, Microliter<\/li>\r\n \t<li>Micropipette<\/li>\r\n \t<li>Accuracy<\/li>\r\n \t<li>Precision<\/li>\r\n \t<li><span data-preserver-spaces=\"true\">Gravimetric method<\/span><\/li>\r\n<\/ul>\r\n<\/div>\r\n<\/div>\r\n&nbsp;\r\n<div class=\"textbox textbox--learning-objectives\"><header class=\"textbox__header\">\r\n<h4 class=\"textbox__title\">Objectives<\/h4>\r\n<\/header>\r\n<div class=\"textbox__content\">\r\n<ul>\r\n \t<li>Review available types of micropipettes.<\/li>\r\n \t<li>To determine which micropipettes\u2019 are best used to measure specific volumes.<\/li>\r\n \t<li>To precisely measure using various sized micropipettes\u2019 (P-20, P-200 and P-1000).<\/li>\r\n \t<li>To accurately prepare samples using each size micropipette.<\/li>\r\n<\/ul>\r\n<\/div>\r\n<\/div>\r\n&nbsp;\r\n<div class=\"textbox textbox--examples\"><header class=\"textbox__header\">\r\n<h4 class=\"textbox__title\">Materials<\/h4>\r\n<\/header>&nbsp;\r\n<ul>\r\n \t<li>Microcentrifuge tubes, 1.5 ml (4)<\/li>\r\n \t<li>Microcentrifuge tube rack<\/li>\r\n \t<li>Micropipettes and tips (P-20, P-200, P-1000)<\/li>\r\n \t<li>Microcentrifuge<\/li>\r\n \t<li>Permanent Marker<\/li>\r\n \t<li>Blue Dye<\/li>\r\n \t<li>Red Dye<\/li>\r\n \t<li>Green Dye<\/li>\r\n \t<li>Yellow Dye<\/li>\r\n \t<li>Wax paper (3)<\/li>\r\n \t<li>Beakers (4)<\/li>\r\n<\/ul>\r\n&nbsp;\r\n\r\n<em>*Colored dyes are stock solutions of 1 ml food coloring and 499 ml of water<\/em><strong><em>.<\/em><\/strong>\r\n\r\n&nbsp;\r\n\r\n<\/div>\r\n<div class=\"textbox shaded\">\r\n<h5>Pre-assessment<\/h5>\r\n<ul>\r\n \t<li>Complete the following conversions:<\/li>\r\n<\/ul>\r\n<ol>\r\n \t<li style=\"list-style-type: none;\">\r\n<ol>\r\n \t<li>500 uL =\u00a0 __________mL<\/li>\r\n \t<li>25 mL = \u00a0___________uL<\/li>\r\n \t<li>2290 uL =\u00a0 __________nL<\/li>\r\n \t<li>875 mL = ___________uL<\/li>\r\n \t<li>3 uL =\u00a0 ____________mL<\/li>\r\n<\/ol>\r\n<\/li>\r\n<\/ol>\r\n<ul>\r\n \t<li>175 uL sample is needed for an experiment. Which micropipette will measure this amount.<\/li>\r\n \t<li>1650 uL sample is needed for an experiment. Which micropipette will measure this amount.<\/li>\r\n \t<li>What do you think would happen to the accuracy of your measurement if the plunger is depressed to the second stop instead of only the first stop when taking up a sample?<\/li>\r\n \t<li>What volume is the pipette set for in the 3 pipettes?<\/li>\r\n<\/ul>\r\n<img class=\"aligncenter wp-image-370 size-medium\" src=\"http:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Picturemicropipette-300x234.png\" alt=\"This photo questions micropipette reading, P20, P200, P1000.\" width=\"300\" height=\"234\" \/>\r\n\r\n<\/div>\r\n&nbsp;\r\n<div class=\"textbox textbox--exercises\"><header class=\"textbox__header\">\r\n<h4 class=\"textbox__title\">Exercise 1: Pipetting Exercise<\/h4>\r\n<\/header>\r\n<div class=\"textbox__content\">\r\n\r\nPROCEDURE\r\n<div>\r\n<ol>\r\n \t<li>Using a permanent marker, label the empty microcentrifuge tubes A, B, C and D.<\/li>\r\n \t<li>Using the smallest micropipette possible, measure the dye solutions into each tube according to Table 1.\r\n<ul>\r\n \t<li>Use a new pipet tip each time a new color dye is used.<\/li>\r\n \t<li>Add the smallest volume first.<\/li>\r\n \t<li>Add the next volume to the inside of the tube without letting the drops touch. The drop will stick because of adhesion.<\/li>\r\n \t<li>When all volumes have been added, bring the drops to the bottom of the tube with a quick wrist flick.<\/li>\r\n<\/ul>\r\n<\/li>\r\n \t<li>Spin tubes A and B in the microcentrifuge for 1 to 2 seconds to aspirate the solutions. Move the tubes to the rack.\r\n<table class=\"aligncenter\" style=\"height: 260px; width: 446px;\" width=\"446\"><caption><strong>Table for Mixing the Solutions<\/strong><\/caption>\r\n<thead>\r\n<tr>\r\n<th style=\"width: 115.104px; text-align: center;\"><strong>Centrifuge Tube<\/strong><\/th>\r\n<th style=\"width: 55.5521px; text-align: center;\"><strong>Red Dye<\/strong><\/th>\r\n<th style=\"width: 59.875px; text-align: center;\"><strong>Blue Dye<\/strong><\/th>\r\n<th style=\"width: 73.2083px; text-align: center;\"><strong>Green Dye<\/strong><\/th>\r\n<th style=\"width: 76.1354px; text-align: center;\"><strong>Yellow Dye<\/strong><\/th>\r\n<\/tr>\r\n<\/thead>\r\n<tbody>\r\n<tr>\r\n<td style=\"width: 115.104px; text-align: center;\"><strong>A<\/strong><\/td>\r\n<td style=\"width: 55.5521px; text-align: center;\">4.0 \u00b5l<\/td>\r\n<td style=\"width: 59.875px; text-align: center;\">5.4 \u00b5l<\/td>\r\n<td style=\"width: 73.2083px; text-align: center;\">2.2 \u00b5l<\/td>\r\n<td style=\"width: 76.1354px; text-align: center;\">-----<\/td>\r\n<\/tr>\r\n<tr>\r\n<td style=\"width: 115.104px; text-align: center;\"><strong>B<\/strong><\/td>\r\n<td style=\"width: 55.5521px; text-align: center;\">6.5 \u00b5l<\/td>\r\n<td style=\"width: 59.875px; text-align: center;\">2.5 \u00b5l<\/td>\r\n<td style=\"width: 73.2083px; text-align: center;\">-----<\/td>\r\n<td style=\"width: 76.1354px; text-align: center;\">2.6 \u00b5l<\/td>\r\n<\/tr>\r\n<tr>\r\n<td style=\"width: 115.104px; text-align: center;\"><strong>C<\/strong><\/td>\r\n<td style=\"width: 55.5521px; text-align: center;\">22.0 \u00b5l<\/td>\r\n<td style=\"width: 59.875px; text-align: center;\">31.5 \u00b5l<\/td>\r\n<td style=\"width: 73.2083px; text-align: center;\">22.0 \u00b5l<\/td>\r\n<td style=\"width: 76.1354px; text-align: center;\">44.5 \u00b5l<\/td>\r\n<\/tr>\r\n<tr>\r\n<td style=\"width: 115.104px; text-align: center;\"><strong>D<\/strong><\/td>\r\n<td style=\"width: 55.5521px; text-align: center;\">-----<\/td>\r\n<td style=\"width: 59.875px; text-align: center;\">210 \u00b5l<\/td>\r\n<td style=\"width: 73.2083px; text-align: center;\">205 \u00b5l<\/td>\r\n<td style=\"width: 76.1354px; text-align: center;\">350 \u00b5l<\/td>\r\n<\/tr>\r\n<\/tbody>\r\n<\/table>\r\n<\/li>\r\n \t<li>Spin tube C in the microcentrifuge for 1 to 2 seconds to pool the solution. Be sure to balance the microcentrifuge with a tube containing an equal volume.<\/li>\r\n \t<li>Flick tube D with a finger to mix the contents.<\/li>\r\n \t<li>Compare tubes A through D with the key tubes provided by the instructor\r\n<ul>\r\n \t<li>For each sample tube, check the level of the final volume AND the final color (indicative of accurate measurement and thorough mixing).<\/li>\r\n \t<li>Indicate in the data table whether your sample tubes were accurately prepared for volume and final color.<\/li>\r\n<\/ul>\r\n<\/li>\r\n \t<li>As an additional check of accuracy:\r\n<ul>\r\n \t<li>Set the micropipette to the correct added volume and carefully withdraw all of the fluid in tube A. The contents should fill the tip with no air space at the bottom of the tip and no leftover fluid in the tube. Repeat with tube B. Record whether there was any air space in the tip.<\/li>\r\n \t<li>Set the micropipette to the correct volume and carefully withdraw all the fluid in the tube.<\/li>\r\n \t<li>The contents should just fill the tip with no air space at the bottom and no leftover fluid in the tube. Record whether there was any air space in the tip.<\/li>\r\n \t<li>Set the micropipette to the correct volume and carefully withdraw all the fluid in the tube.<\/li>\r\n \t<li>The contents should just fill the tip with no air space at the bottom and no leftover fluid in the tube. Record whether there was any air space in the tip.<\/li>\r\n<\/ul>\r\n<\/li>\r\n \t<li>Discard the microcentrifuge tubes and pipet tips into the trash. Rinse the beakers and return to storage.<\/li>\r\n<\/ol>\r\n<\/div>\r\nPRECAUTION\r\n<div>\r\n<ol>\r\n \t<li>Never use the micropipette without the tip in place; this could ruin the piston.<\/li>\r\n \t<li>Never let the plunger snap back during withdrawing or expelling fluid; this could damage the piston. To prevent the damage, slowly draw up the liquid into the pipette tip.<\/li>\r\n \t<li>Never rotate the volume adjustor beyond the upper or lower range of the pipette, as stated by the manufacturer.<\/li>\r\n \t<li>Never immerse the barrel of the micropipette in fluid.<\/li>\r\n \t<li>Never reuse a tip that has been used to measure a different reagent.<\/li>\r\n<\/ol>\r\n<h4>DATA TABLE<\/h4>\r\n<table class=\"aligncenter\" style=\"height: 167px;\" width=\"736\"><caption><strong>Data Table for Micropipetting<\/strong><\/caption>\r\n<thead>\r\n<tr style=\"height: 76px;\">\r\n<th style=\"height: 76px; width: 80.9375px;\"><strong>Tube<\/strong><\/th>\r\n<th style=\"height: 76px; width: 92.3125px;\"><strong>Color of Solution<\/strong><\/th>\r\n<th style=\"height: 76px; width: 106.542px;\"><strong>Accuracy of Volume<\/strong>\r\n\r\n<strong>(yes or no)<\/strong><\/th>\r\n<th style=\"height: 76px; width: 73.6875px;\"><strong>Total Volume<\/strong>\r\n\r\n&nbsp;<\/th>\r\n<th style=\"height: 76px; width: 112.365px;\"><strong>Pipet tip had airspace<\/strong>\r\n\r\n<strong>(yes or no)<\/strong><\/th>\r\n<th style=\"height: 76px; width: 134.865px;\"><strong>Final Check by Instructor<\/strong><\/th>\r\n<\/tr>\r\n<\/thead>\r\n<tbody>\r\n<tr style=\"height: 31px;\">\r\n<td style=\"width: 668.833px; text-align: center; height: 31px;\" colspan=\"6\"><strong>\u00a0<\/strong><\/td>\r\n<\/tr>\r\n<tr style=\"height: 15px;\">\r\n<td style=\"height: 15px; width: 80.9375px;\"><strong>A<\/strong><\/td>\r\n<td style=\"height: 15px; width: 92.3125px;\"><\/td>\r\n<td style=\"height: 15px; width: 106.542px;\"><\/td>\r\n<td style=\"height: 15px; width: 73.6875px;\"><\/td>\r\n<td style=\"height: 15px; width: 112.365px;\"><\/td>\r\n<td style=\"height: 15px; width: 134.865px;\"><\/td>\r\n<\/tr>\r\n<tr style=\"height: 15px;\">\r\n<td style=\"height: 15px; width: 80.9375px;\"><strong>B<\/strong><\/td>\r\n<td style=\"height: 15px; width: 92.3125px;\"><\/td>\r\n<td style=\"height: 15px; width: 106.542px;\"><\/td>\r\n<td style=\"height: 15px; width: 73.6875px;\"><\/td>\r\n<td style=\"height: 15px; width: 112.365px;\"><\/td>\r\n<td style=\"height: 15px; width: 134.865px;\"><\/td>\r\n<\/tr>\r\n<tr style=\"height: 15px;\">\r\n<td style=\"height: 15px; width: 80.9375px;\"><strong>C<\/strong><\/td>\r\n<td style=\"height: 15px; width: 92.3125px;\"><\/td>\r\n<td style=\"height: 15px; width: 106.542px;\"><\/td>\r\n<td style=\"height: 15px; width: 73.6875px;\"><\/td>\r\n<td style=\"height: 15px; width: 112.365px;\"><\/td>\r\n<td style=\"height: 15px; width: 134.865px;\"><\/td>\r\n<\/tr>\r\n<tr style=\"height: 15px;\">\r\n<td style=\"height: 15px; width: 80.9375px;\"><strong>D<\/strong><\/td>\r\n<td style=\"height: 15px; width: 92.3125px;\"><\/td>\r\n<td style=\"height: 15px; width: 106.542px;\"><\/td>\r\n<td style=\"height: 15px; width: 73.6875px;\"><\/td>\r\n<td style=\"height: 15px; width: 112.365px;\"><\/td>\r\n<td style=\"height: 15px; width: 134.865px;\"><\/td>\r\n<\/tr>\r\n<\/tbody>\r\n<\/table>\r\n&nbsp;\r\n\r\n<\/div>\r\n<\/div>\r\n<\/div>\r\n<div>\r\n<div class=\"textbox shaded\">\r\n\r\nDATA ANALYSIS &amp; CRITICAL THINKING\r\n<div>\r\n<ol>\r\n \t<li>By what increment of measurement do the following micropipettes increase (0.1\u00b5L, 1\u00b5L, etc.)?\r\n<ul>\r\n \t<li>P-20 _______________<\/li>\r\n \t<li>P-1000 ________________<\/li>\r\n \t<li>P-200 _________________<\/li>\r\n<\/ul>\r\n<\/li>\r\n \t<li>What volumes, in general, do micropipettes measure?<\/li>\r\n \t<li>What volumes do each of the following types of micropipettes measure?\r\n<ul>\r\n \t<li>P-10 ____________<\/li>\r\n \t<li>P-20 ____________<\/li>\r\n \t<li>P-200 ___________<\/li>\r\n \t<li>P-1000 ___________<\/li>\r\n<\/ul>\r\n<\/li>\r\n \t<li>Why should a micropipette be held at a 45\u00b0 angle when obtaining a sample?<\/li>\r\n \t<li>When obtaining a sample, first depress the plunger on the micropipette to which \u201cstop\u201d?<\/li>\r\n \t<li>Why touch the tip to the inside wall of the reaction tube when expelling a sample?<\/li>\r\n \t<li>Why keep the plunger depressed when removing the pipette from the tube?<\/li>\r\n \t<li>Why is it important to change tips for each new reagent?<\/li>\r\n \t<li>Why is it important to keep a micropipette in a vertical position when there is fluid in the tip?<\/li>\r\n<\/ol>\r\n<\/div>\r\n<\/div>\r\n<\/div>\r\n<div class=\"textbox\">\r\n<h4><strong>Licenses and Attributions<\/strong><\/h4>\r\n<a href=\"https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\" rel=\"cc:attributionURL\">Biology I Cellular Processes Laboratory Manual<\/a>\u00a0by\u00a0The authors &amp; Hillsborough Community College\u00a0is licensed under a\u00a0<a href=\"https:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/\" rel=\"license\">Creative Commons Attribution-NonCommercial 4.0 International License.<\/a>\r\n\r\n&nbsp;\r\n\r\n<\/div>\r\n<div><\/div>\r\n&nbsp;","rendered":"<h2>Volume Measurements using Micropipettes<\/h2>\n<h3>BACKGROUND<\/h3>\n<p>Very tiny amounts of chemicals and biological reagents are used in many molecular biology experiments. To measure these minute <button class=\"glossary-term\" aria-describedby=\"31-334\">volumes<\/button>, technicians use <button class=\"glossary-term\" aria-describedby=\"31-528\">micropipettes<\/button> that measure in <button class=\"glossary-term\" aria-describedby=\"31-348\">microliter<\/button> (\u00b5l) amounts. This <button class=\"glossary-term\" aria-describedby=\"31-515\">metrology<\/button> activity introduces micropipetting technique. As with all fine motor skills, learning how to use a micropipette takes practice and determination. Students must be able to measure these very tiny volumes with <button class=\"glossary-term\" aria-describedby=\"31-520\">accuracy<\/button> and <button class=\"glossary-term\" aria-describedby=\"31-524\">precision<\/button>.<\/p>\n<p>Micropipettes are used to dispense solutions in the <button class=\"glossary-term\" aria-describedby=\"31-348\">microliter<\/button> range. The microliter is a small volume, yet measurable in a typical laboratory. A <button class=\"glossary-term\" aria-describedby=\"31-534\">milliliter<\/button> is a metric unit of volume equal to a thousandth of a liter. A microliter is a unit of volume in the Metric System. The symbol for microliter is \u03bcL. There are 1,000 microliters in a <button class=\"glossary-term\" aria-describedby=\"31-534\">milliliter<\/button>. An example of when you might use microliter volumes would be in preparation of an electrophoresis sample, when isolating DNA, or during chemical purification.<\/p>\n<p>Volume smaller than microliter is <button class=\"glossary-term\" aria-describedby=\"31-352\">nanoliter<\/button>. The prefix nano means one billionth and is symbolized as n in the <button class=\"glossary-term\" aria-describedby=\"31-537\">Metric System<\/button>. Nano denotes a factor of one billionth (10<sup>-9<\/sup>) which means that there are 1,000,000,000 nanoliters in a <button class=\"glossary-term\" aria-describedby=\"31-517\"> liter<\/button>.<\/p>\n<p>Use the following information to calculate metric volume conversions:<\/p>\n<ul>\n<li>1 \u00b5L = 10<sup>\u20136<\/sup> L = 10<sup>\u20133<\/sup> mL<\/li>\n<li>1 nL = 10<sup>\u20139<\/sup><span style=\"font-size: 1em;\"> L = 10<\/span><sup>\u20136<\/sup><span style=\"font-size: 1em;\"> mL<\/span><\/li>\n<li>1000 nL = 1 \u00b5L<\/li>\n<li>0.001 \u00b5L = 1 nL<\/li>\n<\/ul>\n<p><button class=\"glossary-term\" aria-describedby=\"31-528\">Micropipettes<\/button> are among the<span class=\"ls0\">\u00a0essential laboratory instrument used <\/span>to accurately distribute different volumes of liquids in the microliter range. \u00a0In order to prevent cross contamination<span class=\"ls3 ws1\">, m<span class=\"ls0\">icropipettes use disposable tips to aspirate liquid and distribute<\/span><\/span> the solution.Micropipettes enable sterile and accurate liquid handling.<\/p>\n<p>There are several\u00a0types of micropipettes\u00a0on the market which can be classified according to:<\/p>\n<ul>\n<li>Volume: fixed or variable volume\u00a0micropipette<\/li>\n<li>The principle of operation: air displacement or positive displacement\u00a0micropipette.<\/li>\n<li>The operating mechanism: mechanical or electronic micropipette<\/li>\n<li>The number of channels: single-channel or multi-channel\u00a0micropipette.<\/li>\n<\/ul>\n<p>The different parts of a variable volume micropipette are:<\/p>\n<ul>\n<li>Plunger that is used for volume adjustment and to aspirate and dispense the desired amount of liquid into the tip of the micropipette.<\/li>\n<li>Trip Ejector that helps to remove the tip from the micropipette, without touching them, by pressing the tip eject button.<\/li>\n<li>Volume Window that displays the adjusted volume in the volume window (volume set to be aspirated\/dispensed).<\/li>\n<li>Micropipette Shaft is a tube filled with air. Pressing the piston expels a volume of air contained in the rod, releasing the piston allows this air to return to the rod.<\/li>\n<li>Micropipette Tips are tips attached to a micropipette to collect liquid and then transfer it from one place to another. Tips of different sizes are used to collect different volumes of liquid.<\/li>\n<\/ul>\n<figure id=\"attachment_343\" aria-describedby=\"caption-attachment-343\" style=\"width: 1024px\" class=\"wp-caption aligncenter\"><img loading=\"lazy\" decoding=\"async\" class=\"wp-image-343 size-large\" src=\"http:\/\/pressbooks.hcfl.edu\/bsc2010l\/wp-content\/uploads\/sites\/22\/2023\/03\/P1000-1024x576.png\" alt=\"This is a P -1000 micropipette illustrating the different parts of the instrument. Long description is included in the text.\" width=\"1024\" height=\"576\" srcset=\"https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P1000-1024x576.png 1024w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P1000-300x169.png 300w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P1000-768x432.png 768w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P1000-65x37.png 65w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P1000-225x127.png 225w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P1000-350x197.png 350w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P1000.png 1280w\" sizes=\"auto, (max-width: 1024px) 100vw, 1024px\" \/><figcaption id=\"caption-attachment-343\" class=\"wp-caption-text\">P-1000 Micropipette<\/figcaption><\/figure>\n<figure id=\"attachment_341\" aria-describedby=\"caption-attachment-341\" style=\"width: 1024px\" class=\"wp-caption aligncenter\"><img loading=\"lazy\" decoding=\"async\" class=\"wp-image-341 size-large\" src=\"http:\/\/pressbooks.hcfl.edu\/bsc2010l\/wp-content\/uploads\/sites\/22\/2023\/03\/P200-1024x576.png\" alt=\"This is a P- 200 micropipette illustrating the different parts of the instrument. Long description is included in the text.\" width=\"1024\" height=\"576\" srcset=\"https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P200-1024x576.png 1024w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P200-300x169.png 300w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P200-768x432.png 768w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P200-65x37.png 65w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P200-225x127.png 225w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P200-350x197.png 350w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P200.png 1280w\" sizes=\"auto, (max-width: 1024px) 100vw, 1024px\" \/><figcaption id=\"caption-attachment-341\" class=\"wp-caption-text\">P-200 Micropipette<\/figcaption><\/figure>\n<figure id=\"attachment_339\" aria-describedby=\"caption-attachment-339\" style=\"width: 1024px\" class=\"wp-caption aligncenter\"><img loading=\"lazy\" decoding=\"async\" class=\"wp-image-339 size-large\" src=\"http:\/\/pressbooks.hcfl.edu\/bsc2010l\/wp-content\/uploads\/sites\/22\/2023\/03\/P20-1024x576.png\" alt=\"This is a P-20 micropipette illustrating the different parts of the instrument. Long description is included in the text.\" width=\"1024\" height=\"576\" srcset=\"https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P20-1024x576.png 1024w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P20-300x169.png 300w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P20-768x432.png 768w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P20-65x37.png 65w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P20-225x127.png 225w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P20-350x197.png 350w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/P20.png 1280w\" sizes=\"auto, (max-width: 1024px) 100vw, 1024px\" \/><figcaption id=\"caption-attachment-339\" class=\"wp-caption-text\">P-20 Micropipette<\/figcaption><\/figure>\n<p>In a variable volume micropipette the volume of liquid to be aspirated or dispensed are adjusted according to the capacity of the micropipette (a specific minimum and maximum volume range). The acronyms P10, P20, P1000 on micropipettes\u00a0represent the maximum volume that can be aspirated\/dispensed<\/p>\n<p>In the lab, the most common micropipettes are:<\/p>\n<ul>\n<li>P-10 measures volumes of 0.5 \u00b5l to 10 \u00b5l in 0.02 \u00b5l increments;<\/li>\n<li>P-20 measures volumes of 2 \u00b5l to 20 \u00b5l in 0.02 \u00b5l increments;<\/li>\n<li>P-200 measures volumes of 20 \u00b5l to 200 \u00b5l in 0.2 \u00b5l increments;<\/li>\n<li>P-1000 measures volumes of 100 \u00b5l to 1000 \u00b5l in 2 \u00b5l increments.<\/li>\n<\/ul>\n<figure id=\"attachment_493\" aria-describedby=\"caption-attachment-493\" style=\"width: 400px\" class=\"wp-caption aligncenter\"><img loading=\"lazy\" decoding=\"async\" class=\"wp-image-493\" src=\"http:\/\/pressbooks.hcfl.edu\/bsc2010l\/wp-content\/uploads\/sites\/22\/2023\/03\/Micopipette_calculation-300x158.png\" alt=\"This photo illustrates micropipetting volume measurements for P-20, P-200 and P-1000. More details are in the text.\" width=\"400\" height=\"210\" srcset=\"https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Micopipette_calculation-300x158.png 300w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Micopipette_calculation-1024x539.png 1024w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Micopipette_calculation-768x404.png 768w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Micopipette_calculation-1536x808.png 1536w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Micopipette_calculation-2048x1077.png 2048w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Micopipette_calculation-65x34.png 65w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Micopipette_calculation-225x118.png 225w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Micopipette_calculation-350x184.png 350w\" sizes=\"auto, (max-width: 400px) 100vw, 400px\" \/><figcaption id=\"caption-attachment-493\" class=\"wp-caption-text\">Measuring Volumes with Different Micropipettes<\/figcaption><\/figure>\n<p>Operate the micropipette slowly and carefully.<\/p>\n<p>Always use the correct pipet for the job. Most labs have four sizes of micropipettes:<\/p>\n<p>Never attempt to set a volume beyond the pipet\u2019s minimum or maximum values. Only set the volume within the range specified for that micropipette.<\/p>\n<div class=\"row justify-content-around\">\n<div class=\"col-sm-7\">\n<table class=\"aligncenter\" style=\"height: 120px;\">\n<caption>Types of Micropipettes and Their Ranges<\/caption>\n<thead>\n<tr style=\"height: 15px;\">\n<th style=\"width: 163.479px; height: 15px; text-align: left;\" scope=\"col\">Acronym<\/th>\n<th style=\"width: 240.604px; height: 15px; text-align: left;\" scope=\"col\">Volume<\/th>\n<\/tr>\n<\/thead>\n<tbody class=\"text-center\">\n<tr style=\"height: 15px;\">\n<td style=\"width: 163.812px; height: 15px;\">P-2<\/td>\n<td style=\"width: 240.938px; height: 15px;\">0.2 \u2013 2 \u00b5l<\/td>\n<\/tr>\n<tr style=\"height: 15px;\">\n<td style=\"width: 163.812px; height: 15px;\">P-10<\/td>\n<td style=\"width: 240.938px; height: 15px;\">1 \u2013 10 \u00b5l<\/td>\n<\/tr>\n<tr style=\"height: 15px;\">\n<td style=\"width: 163.812px; height: 15px;\">P-20<\/td>\n<td style=\"width: 240.938px; height: 15px;\">2 \u2013 20 \u00b5l<\/td>\n<\/tr>\n<tr style=\"height: 15px;\">\n<td style=\"width: 163.812px; height: 15px;\">P-100<\/td>\n<td style=\"width: 240.938px; height: 15px;\">20 \u2013 100 \u00b5l<\/td>\n<\/tr>\n<tr style=\"height: 15px;\">\n<td style=\"width: 163.812px; height: 15px;\">P-200<\/td>\n<td style=\"width: 240.938px; height: 15px;\">20 \u2013 200 \u00b5l<\/td>\n<\/tr>\n<tr style=\"height: 15px;\">\n<td style=\"width: 163.812px; height: 15px;\">P-1000<\/td>\n<td style=\"width: 240.938px; height: 15px;\">100 \u2013 1000 \u00b5l<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n<\/div>\n<\/div>\n<p><span data-preserver-spaces=\"true\">Micropippete performance is evaluated using a\u00a0<\/span><button class=\"glossary-term\" aria-describedby=\"31-539\">gravimetric method<\/button>. <span data-preserver-spaces=\"true\">Gravimetric methods depend on the high <button class=\"glossary-term\" aria-describedby=\"31-520\">accuracy<\/button> and <button class=\"glossary-term\" aria-describedby=\"31-524\">precision <\/button>of analytical balances. During gravimetric testing, a desired volume of purified water is aspirated and dispensed from the device to be checked. The\u00a0<\/span><em><span data-preserver-spaces=\"true\">desired volume is termed the<\/span><\/em><span data-preserver-spaces=\"true\">\u00a0<\/span>&#8220;nominal volume.<span data-preserver-spaces=\"true\">&#8221; The volume <button class=\"glossary-term\" aria-describedby=\"31-513\">measurement<\/button> is then checked by weighing the liquid on a high-quality analytical balance. A calculation is performed to convert the weight of the water to a volume. If the volume-measuring device is accurately operating, then the nominal volume will be identical to the volume calculated from the weight of the water. This process indicates the <button class=\"glossary-term\" aria-describedby=\"31-520\">accuracy<\/button> of the device being tested.<\/span><\/p>\n<p>Overview and demonstration of\u00a0 miciropipetting technique<\/p>\n<ul>\n<li>Visit <a href=\"https:\/\/www.ncbionetwork.org\/educational-resources\/videos\/micropipetting\"><strong>NC BioNetwork Virtual Microscope<\/strong><\/a><\/li>\n<li>Visit <strong><a href=\"https:\/\/florida.pbslearningmedia.org\/resource\/biot11.sci.life.gen.usingmicro\/using-a-micropipette\/\">Florida PBS Learning Media<\/a><\/strong><\/li>\n<\/ul>\n<div class=\"textbox textbox--key-takeaways\">\n<header class=\"textbox__header\">\n<h4 class=\"textbox__title\">Key Terms<\/h4>\n<\/header>\n<div class=\"textbox__content\">\n<ul>\n<li>Metrology<\/li>\n<li>Measurements<\/li>\n<li>Liters, Milliliters, Microliter<\/li>\n<li>Micropipette<\/li>\n<li>Accuracy<\/li>\n<li>Precision<\/li>\n<li><span data-preserver-spaces=\"true\">Gravimetric method<\/span><\/li>\n<\/ul>\n<\/div>\n<\/div>\n<p>&nbsp;<\/p>\n<div class=\"textbox textbox--learning-objectives\">\n<header class=\"textbox__header\">\n<h4 class=\"textbox__title\">Objectives<\/h4>\n<\/header>\n<div class=\"textbox__content\">\n<ul>\n<li>Review available types of micropipettes.<\/li>\n<li>To determine which micropipettes\u2019 are best used to measure specific volumes.<\/li>\n<li>To precisely measure using various sized micropipettes\u2019 (P-20, P-200 and P-1000).<\/li>\n<li>To accurately prepare samples using each size micropipette.<\/li>\n<\/ul>\n<\/div>\n<\/div>\n<p>&nbsp;<\/p>\n<div class=\"textbox textbox--examples\">\n<header class=\"textbox__header\">\n<h4 class=\"textbox__title\">Materials<\/h4>\n<\/header>\n<p>&nbsp;<\/p>\n<ul>\n<li>Microcentrifuge tubes, 1.5 ml (4)<\/li>\n<li>Microcentrifuge tube rack<\/li>\n<li>Micropipettes and tips (P-20, P-200, P-1000)<\/li>\n<li>Microcentrifuge<\/li>\n<li>Permanent Marker<\/li>\n<li>Blue Dye<\/li>\n<li>Red Dye<\/li>\n<li>Green Dye<\/li>\n<li>Yellow Dye<\/li>\n<li>Wax paper (3)<\/li>\n<li>Beakers (4)<\/li>\n<\/ul>\n<p>&nbsp;<\/p>\n<p><em>*Colored dyes are stock solutions of 1 ml food coloring and 499 ml of water<\/em><strong><em>.<\/em><\/strong><\/p>\n<p>&nbsp;<\/p>\n<\/div>\n<div class=\"textbox shaded\">\n<h5>Pre-assessment<\/h5>\n<ul>\n<li>Complete the following conversions:<\/li>\n<\/ul>\n<ol>\n<li style=\"list-style-type: none;\">\n<ol>\n<li>500 uL =\u00a0 __________mL<\/li>\n<li>25 mL = \u00a0___________uL<\/li>\n<li>2290 uL =\u00a0 __________nL<\/li>\n<li>875 mL = ___________uL<\/li>\n<li>3 uL =\u00a0 ____________mL<\/li>\n<\/ol>\n<\/li>\n<\/ol>\n<ul>\n<li>175 uL sample is needed for an experiment. Which micropipette will measure this amount.<\/li>\n<li>1650 uL sample is needed for an experiment. Which micropipette will measure this amount.<\/li>\n<li>What do you think would happen to the accuracy of your measurement if the plunger is depressed to the second stop instead of only the first stop when taking up a sample?<\/li>\n<li>What volume is the pipette set for in the 3 pipettes?<\/li>\n<\/ul>\n<p><img loading=\"lazy\" decoding=\"async\" class=\"aligncenter wp-image-370 size-medium\" src=\"http:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Picturemicropipette-300x234.png\" alt=\"This photo questions micropipette reading, P20, P200, P1000.\" width=\"300\" height=\"234\" srcset=\"https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Picturemicropipette-300x234.png 300w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Picturemicropipette-1024x797.png 1024w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Picturemicropipette-768x598.png 768w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Picturemicropipette-65x51.png 65w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Picturemicropipette-225x175.png 225w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Picturemicropipette-350x272.png 350w, https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\/wp-content\/uploads\/sites\/22\/2023\/03\/Picturemicropipette.png 1173w\" sizes=\"auto, (max-width: 300px) 100vw, 300px\" \/><\/p>\n<\/div>\n<p>&nbsp;<\/p>\n<div class=\"textbox textbox--exercises\">\n<header class=\"textbox__header\">\n<h4 class=\"textbox__title\">Exercise 1: Pipetting Exercise<\/h4>\n<\/header>\n<div class=\"textbox__content\">\n<p>PROCEDURE<\/p>\n<div>\n<ol>\n<li>Using a permanent marker, label the empty microcentrifuge tubes A, B, C and D.<\/li>\n<li>Using the smallest micropipette possible, measure the dye solutions into each tube according to Table 1.\n<ul>\n<li>Use a new pipet tip each time a new color dye is used.<\/li>\n<li>Add the smallest volume first.<\/li>\n<li>Add the next volume to the inside of the tube without letting the drops touch. The drop will stick because of adhesion.<\/li>\n<li>When all volumes have been added, bring the drops to the bottom of the tube with a quick wrist flick.<\/li>\n<\/ul>\n<\/li>\n<li>Spin tubes A and B in the microcentrifuge for 1 to 2 seconds to aspirate the solutions. Move the tubes to the rack.<br \/>\n<table class=\"aligncenter\" style=\"height: 260px; width: 446px; width: 446px;\">\n<caption><strong>Table for Mixing the Solutions<\/strong><\/caption>\n<thead>\n<tr>\n<th style=\"width: 115.104px; text-align: center;\"><strong>Centrifuge Tube<\/strong><\/th>\n<th style=\"width: 55.5521px; text-align: center;\"><strong>Red Dye<\/strong><\/th>\n<th style=\"width: 59.875px; text-align: center;\"><strong>Blue Dye<\/strong><\/th>\n<th style=\"width: 73.2083px; text-align: center;\"><strong>Green Dye<\/strong><\/th>\n<th style=\"width: 76.1354px; text-align: center;\"><strong>Yellow Dye<\/strong><\/th>\n<\/tr>\n<\/thead>\n<tbody>\n<tr>\n<td style=\"width: 115.104px; text-align: center;\"><strong>A<\/strong><\/td>\n<td style=\"width: 55.5521px; text-align: center;\">4.0 \u00b5l<\/td>\n<td style=\"width: 59.875px; text-align: center;\">5.4 \u00b5l<\/td>\n<td style=\"width: 73.2083px; text-align: center;\">2.2 \u00b5l<\/td>\n<td style=\"width: 76.1354px; text-align: center;\">&#8212;&#8211;<\/td>\n<\/tr>\n<tr>\n<td style=\"width: 115.104px; text-align: center;\"><strong>B<\/strong><\/td>\n<td style=\"width: 55.5521px; text-align: center;\">6.5 \u00b5l<\/td>\n<td style=\"width: 59.875px; text-align: center;\">2.5 \u00b5l<\/td>\n<td style=\"width: 73.2083px; text-align: center;\">&#8212;&#8211;<\/td>\n<td style=\"width: 76.1354px; text-align: center;\">2.6 \u00b5l<\/td>\n<\/tr>\n<tr>\n<td style=\"width: 115.104px; text-align: center;\"><strong>C<\/strong><\/td>\n<td style=\"width: 55.5521px; text-align: center;\">22.0 \u00b5l<\/td>\n<td style=\"width: 59.875px; text-align: center;\">31.5 \u00b5l<\/td>\n<td style=\"width: 73.2083px; text-align: center;\">22.0 \u00b5l<\/td>\n<td style=\"width: 76.1354px; text-align: center;\">44.5 \u00b5l<\/td>\n<\/tr>\n<tr>\n<td style=\"width: 115.104px; text-align: center;\"><strong>D<\/strong><\/td>\n<td style=\"width: 55.5521px; text-align: center;\">&#8212;&#8211;<\/td>\n<td style=\"width: 59.875px; text-align: center;\">210 \u00b5l<\/td>\n<td style=\"width: 73.2083px; text-align: center;\">205 \u00b5l<\/td>\n<td style=\"width: 76.1354px; text-align: center;\">350 \u00b5l<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n<\/li>\n<li>Spin tube C in the microcentrifuge for 1 to 2 seconds to pool the solution. Be sure to balance the microcentrifuge with a tube containing an equal volume.<\/li>\n<li>Flick tube D with a finger to mix the contents.<\/li>\n<li>Compare tubes A through D with the key tubes provided by the instructor\n<ul>\n<li>For each sample tube, check the level of the final volume AND the final color (indicative of accurate measurement and thorough mixing).<\/li>\n<li>Indicate in the data table whether your sample tubes were accurately prepared for volume and final color.<\/li>\n<\/ul>\n<\/li>\n<li>As an additional check of accuracy:\n<ul>\n<li>Set the micropipette to the correct added volume and carefully withdraw all of the fluid in tube A. The contents should fill the tip with no air space at the bottom of the tip and no leftover fluid in the tube. Repeat with tube B. Record whether there was any air space in the tip.<\/li>\n<li>Set the micropipette to the correct volume and carefully withdraw all the fluid in the tube.<\/li>\n<li>The contents should just fill the tip with no air space at the bottom and no leftover fluid in the tube. Record whether there was any air space in the tip.<\/li>\n<li>Set the micropipette to the correct volume and carefully withdraw all the fluid in the tube.<\/li>\n<li>The contents should just fill the tip with no air space at the bottom and no leftover fluid in the tube. Record whether there was any air space in the tip.<\/li>\n<\/ul>\n<\/li>\n<li>Discard the microcentrifuge tubes and pipet tips into the trash. Rinse the beakers and return to storage.<\/li>\n<\/ol>\n<\/div>\n<p>PRECAUTION<\/p>\n<div>\n<ol>\n<li>Never use the micropipette without the tip in place; this could ruin the piston.<\/li>\n<li>Never let the plunger snap back during withdrawing or expelling fluid; this could damage the piston. To prevent the damage, slowly draw up the liquid into the pipette tip.<\/li>\n<li>Never rotate the volume adjustor beyond the upper or lower range of the pipette, as stated by the manufacturer.<\/li>\n<li>Never immerse the barrel of the micropipette in fluid.<\/li>\n<li>Never reuse a tip that has been used to measure a different reagent.<\/li>\n<\/ol>\n<h4>DATA TABLE<\/h4>\n<table class=\"aligncenter\" style=\"height: 167px; width: 736px;\">\n<caption><strong>Data Table for Micropipetting<\/strong><\/caption>\n<thead>\n<tr style=\"height: 76px;\">\n<th style=\"height: 76px; width: 80.9375px;\"><strong>Tube<\/strong><\/th>\n<th style=\"height: 76px; width: 92.3125px;\"><strong>Color of Solution<\/strong><\/th>\n<th style=\"height: 76px; width: 106.542px;\"><strong>Accuracy of Volume<\/strong><\/p>\n<p><strong>(yes or no)<\/strong><\/th>\n<th style=\"height: 76px; width: 73.6875px;\"><strong>Total Volume<\/strong><\/p>\n<p>&nbsp;<\/th>\n<th style=\"height: 76px; width: 112.365px;\"><strong>Pipet tip had airspace<\/strong><\/p>\n<p><strong>(yes or no)<\/strong><\/th>\n<th style=\"height: 76px; width: 134.865px;\"><strong>Final Check by Instructor<\/strong><\/th>\n<\/tr>\n<\/thead>\n<tbody>\n<tr style=\"height: 31px;\">\n<td style=\"width: 668.833px; text-align: center; height: 31px;\" colspan=\"6\"><strong>\u00a0<\/strong><\/td>\n<\/tr>\n<tr style=\"height: 15px;\">\n<td style=\"height: 15px; width: 80.9375px;\"><strong>A<\/strong><\/td>\n<td style=\"height: 15px; width: 92.3125px;\"><\/td>\n<td style=\"height: 15px; width: 106.542px;\"><\/td>\n<td style=\"height: 15px; width: 73.6875px;\"><\/td>\n<td style=\"height: 15px; width: 112.365px;\"><\/td>\n<td style=\"height: 15px; width: 134.865px;\"><\/td>\n<\/tr>\n<tr style=\"height: 15px;\">\n<td style=\"height: 15px; width: 80.9375px;\"><strong>B<\/strong><\/td>\n<td style=\"height: 15px; width: 92.3125px;\"><\/td>\n<td style=\"height: 15px; width: 106.542px;\"><\/td>\n<td style=\"height: 15px; width: 73.6875px;\"><\/td>\n<td style=\"height: 15px; width: 112.365px;\"><\/td>\n<td style=\"height: 15px; width: 134.865px;\"><\/td>\n<\/tr>\n<tr style=\"height: 15px;\">\n<td style=\"height: 15px; width: 80.9375px;\"><strong>C<\/strong><\/td>\n<td style=\"height: 15px; width: 92.3125px;\"><\/td>\n<td style=\"height: 15px; width: 106.542px;\"><\/td>\n<td style=\"height: 15px; width: 73.6875px;\"><\/td>\n<td style=\"height: 15px; width: 112.365px;\"><\/td>\n<td style=\"height: 15px; width: 134.865px;\"><\/td>\n<\/tr>\n<tr style=\"height: 15px;\">\n<td style=\"height: 15px; width: 80.9375px;\"><strong>D<\/strong><\/td>\n<td style=\"height: 15px; width: 92.3125px;\"><\/td>\n<td style=\"height: 15px; width: 106.542px;\"><\/td>\n<td style=\"height: 15px; width: 73.6875px;\"><\/td>\n<td style=\"height: 15px; width: 112.365px;\"><\/td>\n<td style=\"height: 15px; width: 134.865px;\"><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n<p>&nbsp;<\/p>\n<\/div>\n<\/div>\n<\/div>\n<div>\n<div class=\"textbox shaded\">\n<p>DATA ANALYSIS &amp; CRITICAL THINKING<\/p>\n<div>\n<ol>\n<li>By what increment of measurement do the following micropipettes increase (0.1\u00b5L, 1\u00b5L, etc.)?\n<ul>\n<li>P-20 _______________<\/li>\n<li>P-1000 ________________<\/li>\n<li>P-200 _________________<\/li>\n<\/ul>\n<\/li>\n<li>What volumes, in general, do micropipettes measure?<\/li>\n<li>What volumes do each of the following types of micropipettes measure?\n<ul>\n<li>P-10 ____________<\/li>\n<li>P-20 ____________<\/li>\n<li>P-200 ___________<\/li>\n<li>P-1000 ___________<\/li>\n<\/ul>\n<\/li>\n<li>Why should a micropipette be held at a 45\u00b0 angle when obtaining a sample?<\/li>\n<li>When obtaining a sample, first depress the plunger on the micropipette to which \u201cstop\u201d?<\/li>\n<li>Why touch the tip to the inside wall of the reaction tube when expelling a sample?<\/li>\n<li>Why keep the plunger depressed when removing the pipette from the tube?<\/li>\n<li>Why is it important to change tips for each new reagent?<\/li>\n<li>Why is it important to keep a micropipette in a vertical position when there is fluid in the tip?<\/li>\n<\/ol>\n<\/div>\n<\/div>\n<\/div>\n<div class=\"textbox\">\n<h4><strong>Licenses and Attributions<\/strong><\/h4>\n<p><a href=\"https:\/\/pressbooks.hcfl.edu\/Bio1LabManual\" rel=\"cc:attributionURL\">Biology I Cellular Processes Laboratory Manual<\/a>\u00a0by\u00a0The authors &amp; Hillsborough Community College\u00a0is licensed under a\u00a0<a href=\"https:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/\" rel=\"license\">Creative Commons Attribution-NonCommercial 4.0 International License.<\/a><\/p>\n<p>&nbsp;<\/p>\n<\/div>\n<div><\/div>\n<p>&nbsp;<\/p>\n<div class=\"glossary\"><div class=\"glossary__tooltip\" id=\"31-334\" hidden><p>The amount of space a substance occupies. Biologists commonly use units of liters (L), milliliters (mL), and microliters (uL) to express volume.<\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-528\" hidden><p>Micropipettes are a device commonly used in laboratories to aspire and dispense liquid, usually down to 0.1 uL<\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-348\" hidden><p>A measure of volume for a liquid, using the metric system. One microliter (\u03bcL) is equal to a millionth of a liter. <\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-515\" hidden><p>Metrology is the science of measurement and its application. Metrology can be divided into three subfields: scientific metrology, applied metrology, and legal metrology.<\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-520\" hidden><p>Accuracy is how close a given set of measurements are to their true value. A measurement system is considered valid if it is both accurate and precise.<\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-524\" hidden><p>Precision is a description of random errors, a measure of statistical variability. A measurement system is considered valid if it is both accurate and precise.<\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-534\" hidden><p>A milliliter is a unit of volume in the Metric System. The symbol for milliliter is mL. There are 0.001 milliliters in a microliter.<\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-352\" hidden><p>The nanoliter is a unit of Volume in the metric system. It is equal to one billionth of a liter in the International System of Units (SI).<\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-537\" hidden><p>It is the decimal measuring system based on the meter, liter, and gram as units of length, capacity, and weight or mass.<\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-517\" hidden><p>A metric unit of capacity, formerly defined as the volume of one kilogram of water under standard conditions, now equal to 1,000 cubic centimeters.<\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-539\" hidden><p>Gravimetric method includes all analytical methods in which the analytical signal is a measurement of mass or a change in mass.<\/p>\n<\/div><div class=\"glossary__tooltip\" id=\"31-513\" hidden><p>The action of finding out the size, weight, force, length, or amount of something. A measurement can also be a unit or system to calculate a standard measure.<\/p>\n<\/div><\/div>","protected":false},"author":49,"menu_order":3,"template":"","meta":{"pb_show_title":"on","pb_short_title":"Very tiny amounts of chemicals and biological reagents are used in many molecular biology experiments. To measure these minute volumes, technicians use micropipettes that measure in microliter (\u00b5l) amounts. 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