Lab Flashcards
what antiseptics/disinfectants disrupt the lipid bilayer
alcohols, soaps, detergents
what antiseptics/disinfectants are oxidizing agent
hydrogen peroxide, halogens, metals, aldehydes, ethylene oxide, benzalkonium chloride
what antiseptics/disinfectants break H bonds
acids and bases
what is the function of pasteurization
heat to reduce number of microbes but not to zero
besides time, what is the main difference between vat, high temperature, and ultra pasteurization
temperature
what are the temperature differences between vat, high temperature, and ultra pasteurization
vat= 145F (63C)
high temp= 161F (72C)
ultra= 280F (138C)
how does an autoclave work
it holds water at 121C and 2x atmospheric pressure for 15 minutes to destroy all microbes including spores
what type of setting is UV radiation used in for sterilization
hospital
how does ionizing/gamma radiation work
creates free radicals to damage DNA
what is gamma radiation used for
to sterilize items that can’t be put through an autoclave (ex: plastics)
what is involved in liquid filtration
sterilized liquids by passing them through a filter with 2 micrometers
when is liquid filtration used
when heating would kill something you wanted to preserve
how does a hemocytometer/Petroff-Hausser Counter with microscopy work to count bacteria
it uses a microscope slide with grids
the volume of each square is known so you can count the number of cells in each square and multiple to determine the number of bacteria in the liquid sample
how does spectrophotometry work
a line is shined though the liquid sample and the amount of light blocked is used to determine the number of bacteria in the sample by comparing it to McFarlands 0.5 standard
what are 2 limitations with spectrophotometry
production of biofilm increases bacteria size so you may be over-counting
can’t be used with samples that are too dark or too opaque
what is the concentration and number of bacteria in McFarlands standard
0.5
1x10^8 bacteria/mL
how many bacterial colonies must be present on a plate to count
30-300
what are 2 limitations to counting bacteria on a plate
biofilm causes bacteria to stick together so each colony may represent more than 1 bacteria
having more than 300 colonies is likely that some colonies represent more than 1 bacteria leading to undercounting
what is the dilution standard for antibody/antigen samples
1:2
in titers, the __ the second number, the more antibody present (higher or lower)
ex: is 1:16 or 1:64 a greater titer
higher
1:64
what causes zeta potential in RBC
sialic acid on the RBC surface creates a net - charge that IgG is too small to overcome, therefore IgM must be used for crosslinking
what antibody does blood typing use
anti-RBC IgM
what is the question asked for the indirect Coomb’s test
is there IgG that binds to the RBC
*used maternal serum
what is the question asked for the direct Coomb’s test
does the RBC have IgG stuck to it
*uses fetal RBC
how does radial immunodiffusion differ from double immunodiffusion
radial determines the amount of antigen in the sample
double immunodiffusion tells us if the antigen is present or not
do immunoassays use monoclonal or polyclonal antibodies
mono
what is ELISA used for
to detect protein/antigen in serum using antibodies
*can be used to determine titer
what does ELISPOT TSpot test for
how many cells dump out the protein of interest
*more spots on plate= more cells that released the protein/cytokine
ex: if T cells make IFN gamma
“is IFN gamma released in response to tuberculosis antigen?”
what is the function of immunohistochemistry
used to access tissue location a protein localizes in
what results does fluorescence activated cell sorting (FACS) give is
percent of cells that express a specific marker and level of expression
(flow cytometry)
what does neutrophil functional assay tell us
if oxidative burst is functioning and NADPH oxidase is made (shift in flow cytometry= working)
what is the function of leukocyte function assay
determine if B and T cells have normal activity by using something you know will activate all B or all T cells
what 2 things can be used to stimulate T cells in leukocyte function assay
PHA
antibodies against CD3 and CD28
what is used to stimulate B cells in leukocyte function assay
antibody against antibody
(ex: antibody against IgG)
how do you measure T cell proliferation
used fluorescence based assay, dye the cytoplasm, stimulate T cells, notice degree of decreasing fluorescence
how would you perform a functional assay for apoptosis
isolate monocytes/T/B cells
treat with PHA to stimulate T cells
culture with IL-2 for T cell proliferation
add anti-FAS antibody to trigger apoptosis
use TUNEL assay to detect fragmented DNA
*undergoing apoptosis= cell will fluoresce
how can you test for T cell maturation
if cells are actively undergoing VDJ recombination in the thymus, the T cell receptor excision circles will be detected in the blood
if they are absent, there is no VDJ recombination= no T cells= can be used to detect SCIDs
what is the function of CH50 test (complement hemolysis 50%)
test if complement is working or if it has been used up
less complement= CH50 closed to 1:2
what is the Cr51 release assay used to test for
CD8+ T cell function
when should modified acid-fast staining be performed
for actinomyces, fungi, and some parasites