serotyping
S dysenteriae is group A with 13 serotypes
S flexneri is group B with 6 serptypes
S boydii is group C with 18 serotypes
S sonnei is group D with 1 serotype
group A is beg for mannitol fermentation
all others groups are pos
group D is late lactose fermenters and ONPG pos
serotyping is done using slide agglutination tests using specific antisera
Wednesday, March 2, 2011
Tuesday, March 1, 2011
this is getting weird
I am posting stuff on immunology and microbiology 3 to a site that already has microbiology1 stuff on it. Is it me or is it getting too busy here? Let me know
The ELEK test
This test is done to determine if Corynebacterium diphtheriae releases toxin. It is performed after the organism is identified as being C diphtheriae.
Add K tellurite and rabbit serum to molten agar. Dispense this agar to a sterile petri dish using aseptic technique. Place a filter paper strip impregnated with antitoxin across the middle of the plate. Allow the filter paper to sink to the bottom of the the plate. Wait till the plate solidifies and is dry.
do a line streak of the test organism, positive and negative controls at right angles to the filter paper. Incubate
Examine for lines of white precipitation at 45 degree angles a small distance away from the filter paper.
If the positive control is adjacent to the test, and the test is positive, you will see a wave like formation. Very very pretty, dont you think??? Nasty bug though
Add K tellurite and rabbit serum to molten agar. Dispense this agar to a sterile petri dish using aseptic technique. Place a filter paper strip impregnated with antitoxin across the middle of the plate. Allow the filter paper to sink to the bottom of the the plate. Wait till the plate solidifies and is dry.
do a line streak of the test organism, positive and negative controls at right angles to the filter paper. Incubate
Examine for lines of white precipitation at 45 degree angles a small distance away from the filter paper.
If the positive control is adjacent to the test, and the test is positive, you will see a wave like formation. Very very pretty, dont you think??? Nasty bug though
why do we wash the RBC?
Antigens are located on the surface of RBC. Ab are found in plasma
when you remove the RBC from the patients specimen, you have to go past the plasma to get to the RBC. Therefore you may remove some plasma as well as the RBC. Any Ag in the plasma will interfere with the expected agglutination
so we wash the RBC to get rid of all traces of plasma, and of Ab.
when you remove the RBC from the patients specimen, you have to go past the plasma to get to the RBC. Therefore you may remove some plasma as well as the RBC. Any Ag in the plasma will interfere with the expected agglutination
so we wash the RBC to get rid of all traces of plasma, and of Ab.
how to prepare a RBC suspension
dispense approx 0.5ml of RBC to a testtube
fill the tube up to 3/4 with saline
parafilm and mix by inverting tube once
centrifuge at 2000rpm or 0.5g for 5 minutes
remove parafilm, remove saline
repeat above steps twice more (total of 3 times)
at end of last centrifugation, remove as much saline without disturbing the RBC
add saline up to 1/2 tube, mix and note the colour
we are looking for a bright red/scarlet colour
if the suspension is too dark, add more saline and examine again
once the scarlet colour is achieved, use the suspension in your test
fill the tube up to 3/4 with saline
parafilm and mix by inverting tube once
centrifuge at 2000rpm or 0.5g for 5 minutes
remove parafilm, remove saline
repeat above steps twice more (total of 3 times)
at end of last centrifugation, remove as much saline without disturbing the RBC
add saline up to 1/2 tube, mix and note the colour
we are looking for a bright red/scarlet colour
if the suspension is too dark, add more saline and examine again
once the scarlet colour is achieved, use the suspension in your test
ABO blood typing, the test
take 5 testtubes and label as A, B, O, a and b.
A, B and O are the forward grouping tubes. Its purpose is to determine the identity of antigens present. Antigens are found on the surface of RBC, so the patients red cells are added to these tubes. The known is the antibodies, each of which is colour coded.
a and b is the reverse typing. Its purpose is to determine the identity of any antibodies present. Ab are found in serum/plasma, so the patients plasma are added to these tubes. The known are the antigens, which are red in colour.
dispense 1 drop of a 3% RBC suspension to tubes A, B,O
Dispense 1 drop of plasma to tubes a, b
add 1 drop of the relevant Ab and Ag to each tube (e.g.antiA to Tube A, etc)
mix well and incubate at room temperature for 15 minutes
examine each tube for agglutination
record your results and determine the ABO blood type
A, B and O are the forward grouping tubes. Its purpose is to determine the identity of antigens present. Antigens are found on the surface of RBC, so the patients red cells are added to these tubes. The known is the antibodies, each of which is colour coded.
a and b is the reverse typing. Its purpose is to determine the identity of any antibodies present. Ab are found in serum/plasma, so the patients plasma are added to these tubes. The known are the antigens, which are red in colour.
dispense 1 drop of a 3% RBC suspension to tubes A, B,O
Dispense 1 drop of plasma to tubes a, b
add 1 drop of the relevant Ab and Ag to each tube (e.g.antiA to Tube A, etc)
mix well and incubate at room temperature for 15 minutes
examine each tube for agglutination
record your results and determine the ABO blood type
ABO blood groups, the basics
This is just one blood type. The ABO blood type consists of both antigens and antibodies. These antibodies are not specific to the antigens.Ag and Ab are inherited as a set. There are only 2 Ag, antigen A and antigen B. The name of the blood type is taken from the antigen present in the blood. So blood type A has A antigens and antibody B. Blood type B has antigen B and antibody A. Blood type AB has antigens A and B, and no antibodies. Blood type O has no antigens and both antibody A and antibody B.
What I have just described takes place inside the patient, and is said to be in vivo. You need to fully understand this in order to read the agglutination reactions of the blood typing test, and interpret the results.
What I have just described takes place inside the patient, and is said to be in vivo. You need to fully understand this in order to read the agglutination reactions of the blood typing test, and interpret the results.
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