Pipetting Assessment
Learning Outcome #3
- Prepares reagents, standards, and solutions to industry standards for use in the clinical lab, utilizing common lab equipment according to established protocols.
Instructions
Follow the procedural steps in the exact order. Have instructor check off each step.
| Procedural Step | Instructor |
|---|---|
| Put on gloves and eye protection | |
| Assemble equipment and supplies | |
| Check the pipette for any cracks or broken pieces | |
| Hold the pipette lightly between the thumb and last three fingers, leaving the index finger free. | |
| Place the tip of the pipette well below the surface of the liquid to be pipetted. | |
| Using the bulb, carefully draw the liquid up into the pipette until the level of liquid is well above the calibration mark. | |
| Quickly cover the suction opening at the top of the pipette with the index finger. | |
| Wipe the outside of the pipette dry with a piece of gauze or tissue to remove the excess fluid. | |
| Hold the pipette in a vertical position, with the delivery tip against the inside of the original vessel. Carefully allow the liquid in the pipette to drain by gravity until the bottom of the meniscus is exactly at the calibration mark. Do not entirely remove the index finger from the suction-hole end of the pipette; roll the finger slightly over the opening, allow slow drainage to take place. | |
| While still holding the pipette in a vertical position, touch the tip of the pipette to the inside wall of the receiving vessel. Remove the index finger from the top of the pipette to permit free drainage. | |
| Volumetric – Do not blow out Serological – Blow out Mohr – Do not blow out |
|
| Remove the pipette from the receiving vessel and put in pipette washer. | |
| Discard used kimwipes and other contaminated supplies into the bench top biohazard container. | |
| Put glassware in the basin with soapy water. | |
| Clean work area with disinfectant solution. |
Pipetting Exercise #1
Equipment
- 5.0 ml volumetric, 5 ml and 10 ml measuring (Mohr), 5 ml serological pipe! and two glass Pasteur pipets, Pasteur pipe bulb
- Safely bulb, two 100 ml beakers (1 beaker with blue water and 1 empty
beaker), 10 (16 x 100 mm) test tubes, 3 (12×75 mm) test tubes, distilled
water
- Practice transferring 5 ml of blue colored water with a 5 ml volumetric pipe! into a small beaker.
- Practice transferring 5 ml of blue colored water with a 5 ml serological pipe! into the same small beaker.
- Practice transferring 10 ml of blue colored water with a 10 ml Mohr pipe! into the same beaker.
Have an Instructor verify your competency.
| Pipetting Practice | #1 5 ml Volumetric |
#2 5 ml Serological |
#3 10 ml Mohr |
|---|---|---|---|
| Wipe lip each lime before adjusting level | |||
| Hold pipette vertical and al eye level | |||
| Tip against vessel when adjusting initial level. | |||
| Adjust meniscus so it is aligned with calibration mark | |||
| Tip against receiving vessel when draining (note in box y/n) | |||
| Correct delivery: | |||
| Blow-out or not (note in box y/n) | |||
| Correct volume delivered | |||
| Verified by instructor | |||
- Using a 5ml measuring (Mohr) pipet, deliver 1 mL volume of blue/green colored water into each of five (16 x 100mm) test tubes.
- Using a 10 mL measuring (Mohr) piper, deliver 2mL volumes of blue/green colored water into each of five (16 x 100mm) test tubes.
- Using a glass Pasteur pipet, deliver 20 drops of deionized, distilled water and 5 drops of red water to each of 3 (12×75 mm) test tubes using a different angle each time:
- Vertical
- 45°
- Horizontal
Have an instructor verify your work
| Steps Completed Adequately | Initial | |
|---|---|---|
| #4 | Accurately pipetted five 1 mL volumes (5 mL measuring) | |
| #5 | Accurately pipetted five 2 mL volumes (10 mL measuring) | |
| #6 | Delivered three different sized drops with a Pasteur Pipet | |
| Correlated drop size to angle of deliver. (y/n) |
Diluting Pipetting Exercise
| Equipment | Measuring (Mohr or Serological) | 1 mL 2 mL 5mL |
1 2 1 |
|---|---|---|---|
| Volumetric | 3 mL | 1 | |
| Pipets | 16 x 100 | 10 | 1 |
| Test tubes | 1 | ||
| Safety bulb | 1 | ||
| Kim wipes | 1 box | ||
| Parafilm | 10 cut pieces |
Making Dilution
- Set up ten labeled (1-10) test tubes (16 x 100) in numerical order from left to right, in a test tube rack.
- Measure the following solutions into the appropriately labeled test tubes. Pipet the solvent first, then the solute.
| Tube # | Pipet | Water (mL) | Pipet | Colored Solution | ||
|---|---|---|---|---|---|---|
| Size (mL) | Type | Size (mL) | Type | |||
| 1, 2 | 5 mL | measuring | 4.5 mL | 1.0 mL | measuring | 0.5 |
| Cover tubes with parafilm, mix well by inversion | ||||||
| 3, 4 | 2 mL | measuring | 2.0 mL | 2.0 mL | measuring | 2.0 |
| Cover tubes with parafilm, mix well by inversion | ||||||
| 5, 6 | 2 mL | measuring | 1.0 mL | 2.0 mL | measuring | 1.0 |
| Cover tubes with parafilm, mix well by inversion | ||||||
| 7, 8 | 5 mL | measuring | 3.0 mL | 3.0 mL | volumetric | 3.0 |
| Cover tubes with parafilm, mix well by inversion | ||||||
| 9, 10 | 5 mL | measuring | 4.0 mL | 1.0 mL | measuring | 0.5 |