Micropipette Practice Exercise

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Questions and Answers

What should you do if the micropipette's volume knobs are forced past their limits?

This is the correct procedure for calibration.

It is safe to adjust them as needed.

No action is necessary; they are designed for this.

You might damage the micropipette. (correct)

What is the first step to using a micropipette for accurate measurement?

Start distributing the liquid immediately.

Choose a tip that is too large to ensure a good fit.

Always hold the micropipette upside down.

Adjust the volume to the desired amount. (correct)

How should you check if you have pipetted the correct volume into the 1.5 mL tube?

Weigh the tube and its contents.

Confirm with another student.

Self-check using the graduation marks on the tube. (correct)

Estimate visually by looking at the tube.

Which of the following is NOT a correct practice when using a micropipette?

Keeping the micropipette lying down on the table. Signup and view all the answers

What volume should you practice pipetting immediately after confirming comfort with the P-1000?

150 μL Signup and view all the answers

What happens after you press the plunger to the first stop and immerse the tip into the solution?

You must release the plunger slowly to draw the solution in. Signup and view all the answers

What should you do with the used 1.5 mL tube after completing the exercise?

Throw it away into regular waste. Signup and view all the answers

Which step is performed after obtaining a new 1.5 mL microcentrifuge tube?

Pipette 500 μL of colored water into the tube. Signup and view all the answers

What should be labeled on the lids of the 1.5 mL microcentrifuge tubes?

A, B, and C for identification Signup and view all the answers

What is the first step to follow when preparing the reagents for Tube A?

Add the largest volume of any solution first Signup and view all the answers

After mixing the contents of Tube A, what should you do next?

Perform a short spin in a centrifuge Signup and view all the answers

What does a properly pipetted sample in Tube A indicate?

The volume should precisely match 200 μL without leaving solution Signup and view all the answers

How should the accuracy of pipetting be double-checked for Tubes B and C?

Measure the volumes with a micropipette Signup and view all the answers

What should happen to the 1.5 mL tubes after they have been used?

They must be disposed of in regular waste Signup and view all the answers

What is the total volume of solutions combined in Tube B?

50 μL Signup and view all the answers

Which of the following is a common use of absorbance spectrophotometry in laboratory procedures?

To determine the quantity and quality of nucleic acids Signup and view all the answers

What should be considered to avoid introducing air bubbles while pipetting?

Immerse the tip until the plunger returns to the home position. Signup and view all the answers

What is the optimal method for dispensing the solution from a pipette tip?

Aim the tip gently against the wall of the container. Signup and view all the answers

What should be done with the pipette tip after pipetting different solutions?

You can keep it if pipetting the same solution into a clean container. Signup and view all the answers

When mixing reagents in a 1.5 mL tube, which reagent should be added first?

Reagent with the largest volume to facilitate mixing. Signup and view all the answers

What technique is recommended for mixing reagents in a microcentrifuge tube?

Gently flick the tube with your fingers and/or use a vortex. Signup and view all the answers

What is the purpose of centrifuging reagents in a microcentrifuge tube after mixing?

To collect all solution to the bottom of the tube. Signup and view all the answers

Which statement reflects a common misconception regarding pipetting accuracy?

A well-calibrated micropipette guarantees accurate measurements. Signup and view all the answers

How should the end of the pipette tip be positioned when inserting it into a solution?

Kept close to the surface to minimize solution sticking. Signup and view all the answers

What is the relation of OD260 of 1 for double-stranded DNA in terms of concentration?

50 µg/mL Signup and view all the answers

What is the OD260/OD280 ratio indicative of a pure dsDNA solution?

1.8 Signup and view all the answers

Which wavelength is primarily used to assess the concentration of nucleic acids?

260 nm Signup and view all the answers

If the OD260 of a dsDNA sample is 0.70, what is the concentration of the sample?

35 µg/mL Signup and view all the answers

What does an OD260/OD280 ratio smaller than 1.8 generally suggest?

Contamination by proteins or phenol Signup and view all the answers

What is the result of dividing OD260 of 0.70 by OD280 of 0.39 for a dsDNA solution?

1.79 Signup and view all the answers

Which measurement can indicate contamination by urea in nucleic acid solutions?

OD230 Signup and view all the answers

What is the purpose of serially diluting a DNA solution before analysis?

To bring the concentration within measurable limits Signup and view all the answers

What is the purpose of the 'Touch' mode on the benchtop vortex?

To activate the vortex only when samples are pressed. Signup and view all the answers

Which dial is used to control the duration of centrifugation in a microcentrifuge?

Time dial Signup and view all the answers

What should be done after placing samples in the rotor of a microcentrifuge?

Ensure the rotor cover is securely placed back. Signup and view all the answers

How does the rpm/rcf dial affect the operation of a microcentrifuge?

It allows selection between RPM and RCF units for centrifugal force. Signup and view all the answers

What is the maximum speed behavior of the microcentrifuge when the 'short' button is held?

The rotor spins at a maximum speed until the button is released. Signup and view all the answers

What does the term 'rcf' refer to in the context of centrifugation?

Relative Centrifuge Force. Signup and view all the answers

Why is it important to operate a microcentrifuge with balanced samples?

To prevent the centrifuge from shaking or becoming damaged. Signup and view all the answers

What should be done if unusual noises or shaking are heard during operation?

Press the start/stop button to stop the centrifuge. Signup and view all the answers

Study Notes

Micropipetting Technique

Micropipettes have a plunger with two stops; the first provides gentle resistance, and the second dispenses the full volume.
Practice using the micropipette with colored water in 1.5 mL microcentrifuge tubes before performing experiments.
Confirm pipetting accuracy with graduation marks on the tube and get TA approval before proceeding.

Micropipette Usage Guidelines

Use micropipettes carefully, as they are delicate and costly to replace.
Always select the appropriate micropipette based on the volume you are handling, and adjust the knob within its limits.
Attach tips without force; immerse the tip in the solution only after pressing to the first stop.
Release the plunger slowly to avoid introducing air bubbles.
To dispense, gently press the plunger to the first stop, followed by the second stop for complete solution release.

Reagent Mixing Procedure

For setting up reactions, always add the largest volume reagent first in the 1.5 mL microcentrifuge tubes.
Mix reagents by vortexing or flicking the tube after all components are added.
Centrifuge briefly to ensure all liquid is gathered at the bottom of the tube, avoiding reagent loss.

Spectrophotometric Analysis of Nucleic Acids

Use spectrophotometry (OD260) to measure nucleic acid concentration; for dsDNA, 1 OD260 equals 50 µg/mL, while for ssDNA or RNA, it equals 40 µg/mL.
Assess sample purity using OD260/OD280 ratios—values of 1.8 for dsDNA and 2.0 for RNA indicate purity; lower ratios suggest contamination.
Perform serial dilution if the nucleic acid concentration is too high for direct measurement, targeting a final dilution of 200x.

Nanodrop Spectrophotometer Operation

Use a Nanodrop to analyze DNA solutions efficiently with smaller sample volumes.
The device calculates concentration and purity of nucleic acids but requires manual verification of calculations.
When operating, ensure the selector is on 'ON' for continuous motion and 'Touch' for vortex mode.

Microcentrifuge Operations

Use benchtop microcentrifuges for 1.5 mL tube centrifugation to pellet particles and collect solutions.
Load samples in a balanced configuration and secure with a rotor cover.
Control speed (RPM or RCF) and duration of centrifugation to optimize results.
Always monitor for unusual noises or movement during operation and stop the centrifuge if necessary.

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Description

Test your understanding of using a micropipette with this practice exercise. The quiz covers the steps and techniques to properly operate a micropipette, including pressing on the plunger and understanding stops. Make sure to familiarize yourself with the materials and methods before you begin.