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Lecture 5

BIOL 1208 Lecture Notes - Lecture 5: Catechol Oxidase, Catechol, Enzyme Kinetics


Department
Biological Sciences
Course Code
BIOL 1208
Professor
Abel
Lecture
5

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Bio Lab 1208 Final
Lab Techniques (20-25 points)
Pipets
Micropipette (mechanical pipette)
Specifies volume through three digit window
Used for less than or equal to one ml (1000 microliters, L)
PXXX stands for the maximum volume
Volumetric
Designed to deliver a single volume (etched on the side of the pipet)
Draw fluid above the ring and slowly release fluid until the meniscus is exactly at the
ring
Not graduated
Identified by the bulb in the center
Mohr
Graduated but markings stop at baseline before the pipet begins to narrow
The marking near the tip should be the same as the total volume listed at the top of the
pipet
Largest/smallest (max/min) volumes are found near the top of the pipet
Must be accurate both while drawing up and dispensing liquid
Serological
Graduated to deliver. The liquid in the tip counts. Marking continues to the tip.
Largest/smallest (max/min) volumes are listed at the top of the pipet
Appropriate amount of fluid is drawn into the pipet and the entire amount transferred
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Only have to be accurate once when using it
Pasteur
Transfer Pipet
**In transferring solutions, use pipets that can hold the desired volume in order to provide the
best accuracy
Pi-Pumps are used for filling pipets
**Green is for 10ml pipet, blue is for 1ml pipet
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Spectrometers
A spectrometer is an instrument designed to detect the amount of radiant energy absorbed or
transmitted by colored molecules
How it works:
Diffraction grating splits light into its component wavelengths (prism)
Always wipe the cuvette before putting it in the spec because dirt, oil, and grim on fingertips
prevents light from effectively going through the glass, giving erroneous absorbance readings.
Output:
Percent Transmittance (0-100%)
Used to zero and blank
Absorbance (Optical Density Unites from 0-2)
**The relationship between transmittance and absorbance is inversely proportional (for
example, a 0 mM solute would be T=100%, A=0%)
Procedure:
i. Set wavelength to desired number
ii. Set MODE to transmittance
iii. Set transmittance to 0% (sample holder is empty)
iv. Insert blank, set transmittance to 100%
It is important to blank the solution because a spectrophotometer uses the
relative amount of light that passes through a solution to determine its
concentration. Hence, it is important to tell the machine how much light is 100%,
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