MBIO 1010 / Lab 2

How do you smear for an acidic or basic stain?

1. Get a loopful of water on the slide.

2. Get 3–4 loopfuls of culture on the slide.

3. Mix the culture and water into nickel with loop. If cloudy, tens of thousands of cells.

4. Air dry.

5. Heat fix with 3–4 passes over pilot flame.

How do you stain for an acidic or basic stain?

1. Perform acidic or basic smear.

2. Add dye. Let stay for 1 minute.

3. Wash off with gentle water. Blot dry with soft pressure.

Starting from which tube would you get an aliquot from in a dilution series for spread plate and keep the same tip? Why?

Most dilute. If you start from more concentrated to less concentrated with the same tip, you’re concentrating the less concentrated.

Which flame should you use when sterilizing your loop?

The larger blue flame that you get when the Bunsen burner is locked in the down position.

If you need to mix a culture of bacteria in a test tube, what is the best way to do that?

Hold the test tube by the glass, and use a finger on your other hand to tap the tube at the bottom.

How should you sterilize your loop in the microbiology lab?

Hold the loop almost vertically, and place it in a hot blue flame until the entire length of the wire turns red hot for a moment.

When performing a streak plate technique, you should…

Sterilize your loop at the beginning, before dipping it into the culture tube and streaking the first section. You should then re-sterilize your loop before streaking into each new section of the plate. Finally, you should sterilize your loop one last time, before putting it down on the counter.

Where should you place the lid of the Petri plate when you are streaking a plate?

I should not place the lid anywhere. I should hold it right above the plate while I'm streaking and replace it as soon as I can.

What is the fastest way to break the P1000 mechanical pipette?

Turn the dial up past the maximum setting of 1ml (or 1000 μl.)

What does it mean when the box of tips for the mechanical pipettes has a piece of autoclave tape covering the opening?

It means that the box of tips has been sterilized in the autoclave, and has not yet been opened.

What should you do if you are the first person to open a new box of tips for the mechanical pipette?

You should move the piece of autoclave tape from the front of the box to the top, to tell other people that the box has been opened.

How should you discard a pipette tip after you have used it?

You should simply reach over to the nearest sharps container, and depress the tip ejector button so that the tip drops directly into the sharps waste bucket.

To label a test tube, it's best to use a permanent marker and…. where do you do this on the test tube?

Write directly on the surface of the glass (but not on the white frosted area) to make clean-up easy.

After transferring 1 ml from the first test tube, into the next test tube, with a P1000 mechanical pipette, I should…

Discard my pipette tip, place my pipette on the counter, mix the tube by tapping the bottom ten times, and then get a fresh tip to continue my dilution series.

How do you do the actual “spreading” in the spread plate technique?

Hold the hockey stick flat against the surface of the plate, and spin the turntable to spread the sample.

How do you sterilize the glass spreader?

Dip the spreader in alcohol for a few seconds, then try to shake off whatever excess alcohol I can. Finally, I'll give the spreader a quick pass through the flame to burn off any remnants of alcohol.

When staining, you will be reaching across your workstation to use the staining sink in the middle of the bench. When you're reaching across your work station, it is best to…

Have the Bunsen burner turned off entirely. If you need it again after your stain is finished, you can always re-light it later.

Do you need to put a cover slip on your simple stain before it is ready for viewing with the microscope?

No, when the slide is dry, it can be placed on the microscope as it is without a cover slip. To view with the 100x objective, just place a single drop of oil on top of the smear.