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BRL Practical > Lab 1: Lab Skills > Flashcards

Lab 1: Lab Skills Flashcards

1
Q

What gets disposed in tip waste?

A

Gloves, sterile disposable pipets, microfuge tubes, blue and yellow pipet tips.

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2
Q

Where do sharp objects get disposed of?

A

Razor blades go into the red Sharps Containers near lab sinks.

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3
Q

Where does broken glass get disposed of?

A

Report all broken glass to your TA. The waste will go into the blue and white broken Glass Boxes near the door.

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4
Q

Where does Ethidium Bromide waste get disposed of?

A

All EtBr waste should go in the special waste container.

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5
Q

Why is EtBr considered a mutagen?

A

Because it is able to intercalate into DNA. Gloves should be worn at all times when handling EtBr.

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6
Q

Where does chemical waste get disposed of?

A

There are some chemicals you will use this semester that need to be disposed of in a specific waste container. This waste container will be stored in the hood.

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7
Q

Where is bacteria culture waste disposed of?

A

It is harmful to dump this waste down the sink, so you will remove all markings from your tubes and put this water in a specific location. The TA will put this waste into an autoclave before disposing of it.

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8
Q

What is an autoclave?

A

An autoclave is a machine that sterilizes whatever is put into it with high pressure, heat, and/or steam.

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9
Q

Where are bacteria plates and waste disposed of?

A

The bacteria plates will go back to the TA bench and later they will be autoclaved and any gloves that were used to touch the bacteria plates should be thrown out in the medical waste bin.

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10
Q

How do you label a sample during lab?

A

Add your section number, group number, your initials, and a description of what the sample is. (S28-T3-SM-A)

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11
Q

Where do you label a petri dish?

A

Always label the bottom of the dish because lids can be switched.

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12
Q

How are documents labeled?

A

Label documents on the computer with section number-group number-first name last initial- assignment

Ex: S28-T3-StephanieM- Capstone Research

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13
Q

What is an assay?

A

An assay is a tool used in science that helps to quantitatively measure the amount of a single part of a total sample.

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14
Q

Biuret assay- a colorimetric method used to determine the protein content in a solution. Describe the parts that make up the single part and the total sample.

A

The protein is the single part, and the solution is the total sample. We are using the assay to specifically find the protein content, because an assay qualitatively measures the amount of a single part of a total sample.

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15
Q

How does a LabQuest2 and a Vernier work?

A

This system facilitates the collection of data by replacing chemical techniques, providing real time graphical presentations, and saving information directly in an electronic form.

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16
Q

What is Excel used for?

A

Excel simplifies the identification and description of patterns or relationships through its graphing and calculation functions. Computers and associated software facilitate the collection and analysis of data so that patterns and relationships can be discovered, described, and verified.

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17
Q

What is accuracy?

A

Accuracy is how close a measurement is to its true value.

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18
Q

What is precision?

A

Precision is the ability to repeatedly measure a value in a fixed situation and get the same results.

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19
Q

Explain the “target comparison” in terms of accuracy and precision.

A

When an arrow is fired at a bullseye, the closer the arrow is to the bullseye, the more accurate the shot is. If multiple arrows are shot, precision would be the size of the cluster of arrows. A small cluster would indicate high precision, whereas a random scattering would indicate low precision. If many arrows are shot and all hit the bullseye, the shots are both accurate and precise (because they are close to the target and close to each other).

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20
Q

What are the three rules for determining how many significant figures are in a number?

A
  1. Non-zero digits are always significant.
  2. Zeros between two significant digits are also significant.
  3. A final zero or trailing zeros in the decimal portion ONLY are significant.
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21
Q

Significant figures in the lab (When recording data from an instrument), you should…

A

Record all the digits displayed. It is important to use significant figures when recording a measurement so that it does not appear to be more accurate than the equipment is determining. For example, if the scale reads 25.06, record 25.06 and not 25 or 25.0600.

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22
Q

Handling significant figures in calculations

A

Round to 3 significant figures and round numbers only after all of your calculations are complete.

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23
Q

Round to 3 significant figures: 2.3467

A

2.35

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24
Q

Round to 2 significant figures: 0.000429687

A

0.00043

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25
Q

Round to 1 significant figure: 0.00039

A

0.0004

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26
Q

What happens if there is a 5?
If the number before the 5 is odd, __________
If the number before the 5 is even, __________

A

If the number before the 5 is odd, round up.
If the number before the 5 is even, let it be.

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27
Q

Round to 2 significant figures: 2.35
Round to 2 significant figuresL 2.45

A

2.4 (for both)

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28
Q

Metric System Units
(Meaning)
Prefix
mega-
kilo-
milli-
micro-
nano-
pico-

A

mega- One million
kilo- One thousand
milli- One-thousandth
micro- One-millionth
nano- One-billionth
pico- One-trillionth

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29
Q

Metric System Units
(Exponential Notation)
mega-
kilo-
milli-
micro-
nano-
pico-

A

mega- 10^6
kilo- 10^3
milli- 10^-3
micro- 10^-6
nano- 10^-9
pico- 10^-12

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30
Q

Volume
Liter
(Symbol, Molecules, Symbol)

A

Symbol= l (or L)
Molecules= mole
Symbol= mol

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31
Q

Volume
Milliliter
(Symbol, Equivalent, Molecules, Symbol, Equivalent)

A

Symbol= ml
Equivalent= 10^-3L
Molecules= millimole
Symbol= mmol
Equivalent= 10^-3 mol

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32
Q

Volume
Microliter
(Symbol, Equivalent, Molecules, Symbol, Equivalent)

A

Symbol= ul
Equivalent= 10^-6L
Molecules= micromole
Symbol= umol
Equivalent= 10^-6 mol

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33
Q

Nanomole (Molecules)
List Symbol and Equivalent

A

Symbol= nmol
Equivalent= 10^-9 mol

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34
Q

Picomole (Molecules)
List Symbol and Equivalent

A

Symbol= pmol
Equivalent= 10^-12 mol

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35
Q

Mass
Gram
List Symbol

A

g

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36
Q

Mass
Milligram
List Symbol and Equivalent

A

Symbol= mg
Equivalent= 10^-3g

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37
Q

Mass
Microgram
List Symbol and Equivalent

A

Symbol= ug
Equivalent= 10^-6g

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38
Q

1 mL= _________ uL

A

1000 uL

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39
Q

1g= __________ mg

A

1000 mg

40
Q

1mL of water= __________

A

1 gram

41
Q

1000 uL= ___________mL

A

1 mL

42
Q

1000 mg= __________ g

A

1 g

43
Q

1 gram= _____________ mL of water

A

1 mL of water

44
Q

If you need to measure 0.18 mL of liquid, what would the volume be in uL?

A

0.18*1000= 180 uL

45
Q

If you have a mass of 0.25g and want to convert it to mg?

A

0.25g*1000= 250 mg

46
Q

If you want to weigh out 150 mg of a solid on a weighing balance that displays units of grams, how many grams should be displayed on your balance?

A

150mg/1000=0.15g

47
Q

Transfer Pipette

A

You can transfer liquids using a plastic Pasteur transfer pipette. These pipettes have marked gradations on the side and are used to transfer liquids that are less than 1 ml. They are less accurate than using the micropipette.

48
Q

Micropipette P-20, P-200, and P-1000

A

A common practice in laboratory work is to pipette volumes that are in the microliter range (10^-3 milliliters). The success and reproducibility of your experiments will depend upon accurate delivery of these small volumes.

49
Q

Ranges for P-20, P-200, and P-1000

A

P-20 for dispensing 2-20 ul
P-200 for dispensing 20-200 ul
P-1000 for dispensing 100-1000 ul

50
Q

The range for each micropipette is labeled _____________

A

On the top of the plunger.

51
Q

What disposable tips are required for the P-20, P-200, and P-1000?

A

P-20: Yellow tip
P-200: Yellow tip
P-1000: Blue tip (larger)

52
Q

Increments for P-20, P-200, P-1000

A

P-20= increments of 0.1 ul
P-200= increments of 1 ul
P-1000= increments of 10 ul

53
Q

How do you keep pipet tips sterile?

A

Throw them out after use, they are no longer sterile. Once the pipet tip has touched the bench, your hand, or sleeve, it is no longer sterile and should be thrown away.

54
Q

When filling tips:

A
  • Go to first stop before entering the liquid you want to pipet
  • Insert into liquid and slowly draw the liquid into the tip
  • Go slowly especially in viscous liquid, be sure not to touch the bottom
55
Q

When dispensing liquid:

A
  • Move tip to container and press plunger to the first and then second stop, then eject tip into tip waste
  • Always use a fresh tip or pipet
56
Q

Pipet Bulbs (steps for use)

A

Used for mL volumes (1-10mL)
Sterile technique
Steps:
- Squeeze air valve (A) and deflate air from bulb, should stay collapsed
- Place bulb onto siphon, keep vertical at all times, now slowly draw up liquid by squeezing suction valve (S)
- Expel using E valve

57
Q

Percent Error formula

A

Theoretical - Measured Mass
____________________________ x 100
Theoretical Mass

58
Q

The activity of proteins is very dependent on what?

A

Very dependent on pH, salt concentrations, and temperature of the reaction mixture. In some cases, a change in the pH from 7.5 to 6.5 or a 10 degree change in the temperature may cause greater than a 1000-fold reduction in the protein’s activity. It is very important to understand the function of the proteins involved in the experiment to know their optimal conditions for activity.

59
Q

What is a buffer?

A

An aqueous solution containing a specific mixture of salts, buffering agents, sometimes reducing agents, detergents or cofactors, in which each of the components has a purpose and is intended to optimize the reaction. The basic function of a buffer is to resist changes in hydrogen ion concentration. Buffers are extremely important in helping maintain the pH of a solution.

60
Q

What is a stock solution?

A

Saves time and space. These are highly concentrated solutions that last over long periods of time. They take up less space, and these stocks are easily diluted for use when necessary. Stock solutions are usually diluted with water.

61
Q

What is the equation used to make a specific volume of a dilute solution from a stock solution?

A

C1V1=C2V2

62
Q

What are the components that make up the equation C1V1=C2V2?

A

This is the equation used to make a specific volume of a dilute solution from a stock solution.
C1= concentration of stock solution
V1= volume of stock solution needed to make dilute solution
C2= final concentration of dilute solution
V2= final volume of dilute solution

63
Q

What is the dilution factor?

A

The factor by which the concentration of the dilute solution is reduced compared to the concentration of the stock solution. The dilution factor is derived from the equation C1V1=C2V2 and is defined as
DF= C1/C2=V2/V1

64
Q

Explain how to make a 2-fold serial dilution starting from a bromophenol blue stock solution that has a concentration of 8 mg/ml.

A
  1. Obtain 4 cuvettes. Label the tops of the cuvettes 0 through 4 and B for blank with a sharpie.
  2. Add 2 ml of bromophenol blue (8 mg/ml) stock solution to the cuvette labeled “0” using the P-1000 (twice).
  3. Fill each of the cuvettes labeled 1 through 4 with 1 ml of 1X TAE buffer using the P-1000.
  4. Switch the vortexes to “On” and set the dial to 6.
  5. Transfer 1 ml of stock solution (transfer volume) from cuvette 0 to cuvette 1. Put a cuvette cap on the cuvette. Vortex to mix.
  6. Uncap cuvette 1 and transfer 1 ml from cuvette 1 to cuvette 2. Cap and vortex to mix. Repeat this procedure through cuvette 4. Discard the excess 1 ml from cuvette 4 into the reinstate beaker.
  7. Prepare your Blank: add 1ml of 1X TAE buffer to the cuvette labeled B (you are using TAE buffer as the blank for this experiment).
  8. Cap all cuvettes and set aside until you are ready to read the absorbances.
65
Q

Where do the Vernier probes connect to the LabQuest?

A

Via the analog ports (on the opposite side of the power button, there are 3 of them)

66
Q

Where does the SpectroVis connect to the LabQuest?

A

Via the USB port (on the opposite side of the power button, next to the three analog ports)

67
Q

What is the total dilution factor?

A

How diluted the tube is from the stock solution (5X, 25X)

68
Q

What is the step dilution factor?

A

How much each tube is diluted by (all 5X, etc)

69
Q

What is the water volume?

A

How much volume is needed to dilute the sample or stock solution

70
Q

What is the transfer volume?

A

How much volume you have to move (transfer) to the next tube

71
Q

What is the total volume?

A

The volume of the solution/liquid before you proceed to transfer the “volume transferred”

72
Q

What is the meter screen in LabQuest used for?

A

To measure discrete values, calibrate probes, and set up collection parameters.

73
Q

What is the graph screen in LabQuest used for?

A

To monitor data collection, as well as determine simple statistical data information such as the average for a select data sample.

74
Q

What is the table screen in LabQuest used for?

A

To see an overview of your collected data values in table format.

75
Q

How do you access the calculator through LabQuest?

A

Go to the Home screen. Press the Accessories folder, then press Calculator.

76
Q

How do you turn off the LabQuest?

A

Go to the Home screen. Press the System folder, then press Shut Down.

77
Q

What is a SpectroVis?

A

A spectrophotometer.

78
Q

What is a spectrophotometer?

A

A machine that can measure absorbance or transmittance of a pigmented solution. Biologists commonly use them to quantify the concentration of materials in a solution.

79
Q

How does a spectrophotometer work?

A

This instrument produces a beam of light with a specific wavelength that passes through the sample before entering a photometer that measures the amount of light. This measurement is transformed electronically to a reading on a meter quantifying the amount of light absorbed.

80
Q

What is important about the absorbance and the concentration of a sample?

A

The absorbance of a sample is directly proportional to the concentration of material in the sample.

81
Q

The SpectroVis needs to be calibrated using __________ appropriate for each experiment and/or any time it is unplugged from the Lab Quest.

A

The blank

82
Q

The blanks in our procedures ______________ the reagants.

A

Include

83
Q

Cuvettes

A

A cuvette is a small tube used to hold samples for measurements with a spectrophotometer. One side of the cuvette has an arrow at the top of the face that indicates the light path orientation. The cuvette arrow should be oriented toward the arrow on the SpectroVis cuvette holder when placing the cuvette in the SpectroVis. Failure to orient the cuvette in the SpectroVis properly will result in erroneous measurements. The cuvette must be filled with at least 1ml of liquid to obtain an accurate measurement with the SpectroVis. Always check that the level of the liquid is above the 1ml mark on the cuvette. If the liquid is below the 1ml mark, you have pipetted incorrectly and should redo the sample. The widow of the cuvette is where light passes through the sample being measured. Always hold and label the cuvettes at the top half above the 1ml mark to avoid fingerprints and marks in the light path that can affect the absorbance values. Always wipe the outside of the cuvette with a Kimwipe before putting it in the SpectroVis to remove dirt, fingerprints, and liquid. It is extremely important to wipe off any liquid on the outside of the cuvette as it can damage the equipment.

84
Q

Why are standards used?

A

Standards have a KNOWN amount of the material being assayed and are used to calculate the amount of material in the unknowns.

85
Q

What is one way to determine the concentration of an unknown sample?

A

Cs/Cu=As/Au
Cs= concentration of the standard
Cu= concentration of the unknown
As= absorbance of the standard
Au= absorbance of the unknown

So to find the concentration of an unknown, use the equation:
Cu= Au*Cs/As

86
Q

What is the second way to determine the concentration of an unknown sample?

A

Making a standard curve.

87
Q

What is a standard curve?

A

A tool that scientists use to determine the unknown concentration of a sample.

88
Q

How is a standard curve generated?

A

By measuring the absorbance of a series of standards and graphing the absorbance as a function of concentration. The unknown concentration of a sample can be determined by the interpolation on the graph (estimating unknown values that fall between known values) or you can calculate the concentration (x) of unknown samples based on the measured absorbance (y) using the graph’s linear trend line equation.

89
Q

What is linear regression?

A

Linear regression is a statistical method for modeling the relationship between two variables, x and y.

90
Q

What is a linear trendline?

A

A straight line is best fit to the data points by minimizing the deviation of the data points from the line.

91
Q

What is the R-squared (R^2) value?

A

The R^2 value provides a measure of how well the linear trendline fits the data. An R^2 value of 1 indicates a perfect match of the trendline to the data points. An R^2 value of 0 indicates there is no relationship between the values of x and y. If the linear trendline has a good fit, then the trendline equation can be used to calculate one variable from the other (i.e. calculate x based on the value of y and vice versa)

92
Q

If you get an R^2 value of 0.97 or better you can be _________ in your results.

A

Confident. If you get a lower R^2 value, think about some of the steps in the serial dilution or when the cuvettes were measured in the spectrophotometer that could be potential sources of error (equipment, pipetting, clean cuvettes, calibration as well as the accuracy of the person performing the dilution) that can be remedied in the future.

93
Q

What is needed when creating a graph?

A

Title, labels on all axes, and a legend. A key is optional.

94
Q

What is proper graph format?

A

When constructing a graph, the independent variable is plotted on the X-axis while the dependent variable is plotted on the Y-axis. You determine which variable is which by considering which one depends on the value of the other.

95
Q

What is a legend?

A

Every figure must have an explanatory paragraph at its base that reviews the results shown in the figure, the experimental conditions that generated the results, and the statistics. A figure’s legend is an explanatory paragraph that summarizes the content of the figure, not just an identifier for objects on a figure (a key). Be careful, a true legend is not the same thing as what Excel calls a “legend” when you construct a graph.

96
Q

Steps to measure absorbance for a serial dilution (SpectroVis and LabQuest)

A
  • Connect the SpectroVis to the LabQuest via the USB plug.
  • Power on the LabQuest by pressing on the power button on the side. DO NOT press down too hard as this will damage the power button. Wait about 1-2 minutes for the LabQuest to load completely.
  • In the LabQuest app, go to the Meter Screen by pressing it in the upper left corner.
  • Go to Sensors in the menubar at the top of the screen and select Calibrate from the drop-down menu. Wait for 90 seconds for warm up to finish. Place the cuvette labeled “B” (blank) in the cuvette holder of the SpectroVis and press Finish Calibration. When the calibration is complete, press OK.
  • Go to Sensors and select Data Collection. Change the Mode from “Full Spectrum” to “Time Based”. Press OK.
  • Tap on the large red box on the screen and select Change Wavelength from the drop-down menu. Enter the value 550 and press OK. Now you will be reading the absorbance of your solution at one specific wavelength.
  • Remove cuvette B and put cuvette 0 in the cuvette holder of the SpectroVis.
  • To determine the absorbance over time, begin data collection by pressing play. Collect data for at least 10 seconds.
  • When the sampling run is complete, stop data collection.
  • Go to Analyze in the menubar at the top of the screen and select Statistics from the drop-down menu. Select your run. Record the average (mean) absorbance in your table.
  • Remove cuvette 0 and put cuvette 1 in the cuvette holder. Press Play. You will be prompted to either save or discard the data from the previous run before continuing. Press Discard to delete the previous run and start data collection for cuvette 1.
  • Repeat steps 5-8 for all 5 samples in your serial dilution.
  • Now, measure the absorbance of the unknown solutions.
97
Q

What is the equation for calculating the concentration of an unknown solution?

A

Cu= Au*Cs/As

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