Instructions Flashcards
Differentiate between a serological pipette and a Mohr pipette.
- Mohr; some solution remains in after dispensing
- Serological: all solution dispensed for accurate volume; markings all the way to bottom of tip
When pipetting, what volume has the greatest level of error?
Smaller volumes
What type of pipetting should be used for aqueous solutions?
e.g., buffers or diluted salt
Forward
What type of pipetting should be used for viscous solutions?
e.g., protein or glycerol
Reverse, users should pipet slowly to avoid bubbles and inaccuracies
What type of pipetting should be used for volatile compound solutions?
e.g., methanol; hexanes
Reverse
What type of pipetting should be used for nucleotide solutions?
e.g., DNA; PCR products
Forward
What type of pipetting should be used for acids and alkali solutions?
e.g., HCl or NaOH
Forward
What type of pipetting should be used for toxic sample solutions?
Reverse or forward
When should you never reverse pipette?
If the volume exceeds 75% of the maximum pipetting volume; extra liquid will flood the shaft resulting in damage to, or contamination of, the pipette.
Describe forward pipetting.
- Depress to stop 1.
- Aspirate
- Depress to stop 1
- Depress to stop 2 to completely empty tip.
Describe reverse pipetting.
- Depress to stop 2.
- Aspirate
- Depress to stop 1.
- Do not fully empty tip.
- Depress to stop 2 to return excess fluid to original sample or disposal container.
What happens if the reagent changes from green to purple before adding a protein sample?
Biuret reaction a.k.a. BCA protein assay
Glassware or pipette tips contaminated reagent (very sensitive to metal ions)
What are acids and bases according to the Bronsted-Lowry Theory of neutralization?
What defines pH?
- Acids are capable of donating protons.
- Bases are capable of accepting protons.
- pH is the negative logarithm of the activity of hydrogen ions; pH = log (1/[H+]) or pH = -log [H+]
Why does pH change with temperature?
- As temperature rises, molecular vibrations increase which results in the ability of water to ionize and form more hydrogen ions.
- As a result, the pH decreases as temperature increases.
What is a potentiometer?
- pH meter
- Composed of 2 electrodes and a voltmeter
- Hydrogen ion concentratoin (activity) is determined by the voltage that develops between the two electrodes.
- The Nernst equation relates the electrode response to the activity.
How do you prepare BCA reagent from reagents A and B?
50 parts A to 1 part B
How long is prepared BCA working reagent stable?
For a short duration in a closed container at room temperature
What is required of the diluent in microbial enumeration?
- osmolarity simiilar to the sample (otherwise microbes may lyse and die)
- does not support microbial growth
What are two important sources of error in preparing microbial serial dilutions?
How can they be avoided?
- Sampling error resulting from unequal distribution of bacteria in original sample or dilution tubes; avoid by ensuring representative sample and always mix well before removing an aliquot
- Inaccuracy in pipetting; avoid by always examining pipette to ensure correct use, and limit the number of transfers within reason since more transfers = greater chance of error.
What are some factors that may result in a gram positive bacteria staining as if it were gram negative?
- Overheating during heat fixation, over decolorization with alcohol, and even too much washing with water between steps may result in gram-positive bacteria losing the crystal violet-iodine complex
- Cultures more than 24 hours old may lose their ability to retain the crystal violet-iodine complex
- The organism itself; some gram-positive bacteria are better able to retain the crystal violet-iodine complex than others.
Therefore, use precise technique and interpret results with discretion.
Which type of bacteria has a thick peptidoglycan layer?
Gram positive
(no outer membrane)
Which type of bacteria has a thin peptidoglycan layer?
Gram negative
Outer membrane
How is the gas flow rate of a Bunsen burner adjusted?
Gas adjustment knob.
Using the gas supply is poor technique since the adjustment is insensitive.
Describe effective packaging material.
- Nontoxic
- Protects against microbial, insect, and other rodent contamination
- Barrier to moisture, and oxygen
- Protects against environmental toxicants
- Filters out UV light
- Provides resistance to physical damage.
What are the main factors that determine the shelf-life of a food?
- Rate of oxygen and moisture transfer through packaging
Lactic acid is the major product of lactose fermentation.
What are the minor products?
- Acetate
- Ethanol
- Volatile fatty acids
- Contribute to aroma
Why does yogurt thicken during fermentation?
Increasing acidity coagulates milk proteins
What is the best quality indicator for foods that contain weak acids?
Titratable acidity, since it measures undissociated acids in a solution in addition to the [H+]
Why is phenolphthalein referred to as a secondary indicator?
It changes from clear to pink ~8-9.6 pH. This is only close to the equivalent point where the moles of acid equals the moles of base and does not directly identify the equivalence point.
Why is vacuum drying a more accurate method for determining % moisture in a food?
By applying low pressure, water can be removed using lower temperatures and shorter drying times than air-drying.
What is ash?
Inorganic residue that remains from the incineration or complete oxidation of organic matter, and represents the total mineral content in foods.
What may be influenced by manipulation of water activity in food?
- Texture
- Non-enzymatic browning reactions
- Enzymatic activity
- Lipid oxidation
- Microbial growth
What may colour of food indicate?
- Nutrition
- Flavour
- Functional values
- Consumers use colour when deciding what to purchase
What foods are particularly sensitive to moisture and oxygen transfer through packaging?
Foods that are dehydrated or contain appreciable quantities of lipids or other oxygen-sensitive components.
