noelrbrown

HemoBioScience has some "Simulated Patient Plasmas" that work quite well - and "keep" past their expiration.  They even work when aliquoted and frozen.
We have, in the past, requested "Antibody Containing Plasmas" from our blood suppliers.  We use several different suppliers - some will and some won't save them for us.  Usually pay a nominal fee - but not nearly as much as manuf. antisera.  When we get them, we aliquot and freeze.
The least favorite way is  - when a patient has an antibody - or you ID a new antibody - we scavenge as many CBC's as possible, combine the plasmas and freeze.
 
We do have some dilute antisera - but as said before - often times does not work as expected.

Hi @Pamo,
There are a few options.
If you subscribe to a proficiency testing program, you can often save what they send you.  WARNING - do not use these samples until after you have received the results if you are a CLIA lab. You can spike your training samples with commercial antisera.  These don't always react as expected / hoped for. In my opinion, the best option is to save a recent prior patient sample that is no longer needed or expired. Commercial samples are also available, such as this from Hemo bioscience.

Thanks for letting us know, I worked with Imelda on a couple of projects when she was at Liverpool BTS.  I always enjoyed her humour and approach to life.
 
Noel

It is with immense regret that I have to say that I learned yesterday that Dr Patricia Tippett died at the age of 93 on 1st August 2023.

I first met Pat in the early 1970's, when I was a callow youth who had just left school and was working at the IBGRL when it was in Gatliff Road in London (when Dr Kenneth Goldsmith was the Director) and Pat was working in the opposite building in the MRC Blood Group Unit, then under Drs Rob Race and Ruth Sanger.

Pat is probably most famous for her work on the Rh Blood Group System, including categorising the then know Partial D types and for realising that the RH genes were twofold; namely RHD and RHCE.
She was. of course, one of the greats, but was as friendly to this callow youth as I started out in the profession, as she was to everyone who were already greats within the field.

May she rest in peace.

I'm all for the concept of quality and the strive to provide the safest blood products to patients, but I won't deny that sometimes many of our current practices in blood banking in terms of achieving that "quality" seems excessive, unnecessary, and sometimes it feels like a mere quality charade for inspectors and regulators. Considering the hight cost that blood banks have to incur to meet all quality regulations, it may be worth studying the financial impact of the many quality measures that regulate the practice of blood banking and to what extent these measures are actually contributing to achieving the quality needed to provide the best blood products to patients.

Sad news indeed, thanks for letting us know.
 
Noel

If you subscribe to a whole Blood Proficiency test ( like CAP JAT) the samples contain reverse group antibodies anti A, Anti B etc. and a couple of antibodies. its easy to write your own study looking for carryover in the adjacent samples.  Alternatively Hemo bioscience makes and sells a validation kit for instrumentation and this contains similar materials....

Here is an example of how to calculate numbers of units to be screened from a Proficiency test we designed a couple of years ago.  It makes a nice worked example.
 
Frequencies and Donor Screening

The frequency of the c (small) antigen is 80% in Caucasians, 96% in Blacks, and 47% in Asians (Refer to Table 3 for additional information regarding Rh antigen frequencies in various populations).3


 
Antigen Symbol

Caucasian

Black

Asian

D

85%

92%

99%

C

68%

27%

93%

E

29%

22%

39%

c

80%

96%

47%

e

98%

98%

96%

Table 3: Common Rh blood group antigen frequencies3


 
The frequency of the K antigen in 9% in Caucasians and 2% in Blacks. The K antigen can be found at higher frequencies in specific populations—found in approximately 12% of Iranian Jews and can be found in up to 25% of the Arab population (Refer to Table 4 for additional information regarding the antigen frequencies associated with the Kell blood group system).


 
Antigen Symbol

Caucasian

Black

Other

K

9%

2%

Iranian Jews = 12%

Arabs = As high as 25%

k (small)

99.8%

100%


 
Kpa

2%

Less than 0.01%


 
Kpb

100%

100%


 
Jsa

0.01%

20%


 
Jsb

100%

99%


 
Table 4: Common Kell blood group antigen frequencies3


 
Problem #1: How many units of pRBCs should be crossmatched in order to find 3 antigen negative units?


 
1.      Given that ~80% of the population will have the c (small) antigen, ~20% of the donors should be negative for the c (small) antigen. Additionally, given that ~9% of the population will have the K antigen, ~91% of donors should be negative for the K antigen.

**NOTE: When multiple antigen frequencies are involved, you must multiply the antigen negative frequencies together. In this case we would multiple 20% and 91% to account for the donors who are c (small) negative and K negative.

2.      Number of units to be crossmatched = x


   
 
 
 
x = 3 units/0.18 = 16.7 (or 17 units)


 
3.      The blood bank should crossmatch 17 units, where 3 of the 17 units should be compatible.

Thanks for letting us know, I worked with David a long time ago on an IgG quantitation assay.

Depends on the beer . My Belhaven Black better not be ice cold.

Thanks all - that's what I suspected. My patient's antibody is still reacting fairly strong in solid phase, so I'm relying on crossmatch for Cob donors. Think I"ll freeze some plasma for screening purposes in case his titer drops.

Add some Dextran to a plasma sample, it should do the trick and will save you a lot of time and effort freezing and retaining samples etc.

I suggest you try any high potency IgM anti D which these days is a monoclonal and thus not high protein. The anti D on your bench right now ( immediate spin) should work.

I suggest you try any high potency IgM anti D which these days is a monoclonal and thus not high protein. The anti D on your bench right now ( immediate spin) should work.

Add some Dextran to a plasma sample, it should do the trick and will save you a lot of time and effort freezing and retaining samples etc.

So.. just a quick word of warning, its easy to grind up seeds and do an extraction. But what i have found is that the extraction may or may not work and its a bit fickle. if you go down the path of making your own lectin you must QC it by using either polyagglutinable cells or at the very least prepare Neuraminidase treated cells. If you don't you could end up with false negatives.

So.. just a quick word of warning, its easy to grind up seeds and do an extraction. But what i have found is that the extraction may or may not work and its a bit fickle. if you go down the path of making your own lectin you must QC it by using either polyagglutinable cells or at the very least prepare Neuraminidase treated cells. If you don't you could end up with false negatives.

I haven't heard it called Flying squad blood for Donkeys years, srichar3 are you from the UK?

I haven't heard it called Flying squad blood for Donkeys years, srichar3 are you from the UK?

Hemo bioscience sells Polyclonal anti K and Anti Cellano, both are FDA approved potency and could probably be diluted in a 6%BSA solution to be used for this purpose.  info@hemobioscience.com for pricing info...  Also your local American Red Cross may have donated units of Anti Kell plasma you might use but obviously watch out for the ABO types as these are not adsorbed..... 

Malcolm, I agree with you and understand exactly what you are saying.  My challenge is more to do with interpreting requirements from reagent manufacturers and regulatory bodies.  Therefore that is why i said "we" it being the Royal "we".

I would advise people to look at RH/index.htm.  There are now WELL over 100 (almost 200) different weak D types, not to mention the number of Partial D types.  Unless you have access to ALL of these red cells, AND use them as a control EVERY time you perform this test, you cannot QA/QC this test.  There are times, even in the world of blood transfusion/blood group serology, you just have to admit defeat and keep your fingers crossed!  Even if you were using molecular, rather than serological techniques, you would have to perform complete RHD sequencing each time to ensure you would detect all known mutations AND any novel mutations.

Hi Bertie, I'll do my best.
The very best way to adsorb out an auto-antibody is to use autologous red cells (i.e. auto-adsorption). Such an adsorption will take out any auto-antibody, leaving behind any alloantibody, however rare the specificity of the alloantibody may be. To put it another way, if the auto-antibody is panreactive, but underneath that is, say, an anti-Lan (an antibody directed against a very high frequency antigen), however many times you adsorb the patient's plasma with autologous red cells, you will not remove the anti-Lan because the patient is, by definition of the fact that the anti-Lan is an alloantibody, Lan Negative.
BUT, you can only perform an auto-adsorption if you have sufficient red cells (which would be unusual - as a patient with an auto-antibody would normally have a low haemoglobin and haematocrit) and if the patient has not been transfused within the previous 3 months (unlikely, because of the low Hb and hct), because some of the transfused red cells may be present, and may take out the alloantibody.
So, under such circumstances, you would have to go for a differential alloadsorption.
It is important to remember, at this stage, that most (but not all by any means) auto-antibodies are directed against Rh antigens. It is, therefore, vital that you choose three different cells, one of which is R1R1, one of which is R2R2 and one of which is rr. These must be fully typed, so that one is Jk(a+b-), whilst another is Jk(a-b+), one is K+, etc, etc, so that any alloantibody is adsorbed out by one (or two) of the cells, but not by all three.
The red cells used, initially anyway, should be enzyme-treated (to enhance any reaction between an Rh antibody/antigen). At this stage, of course, the MNS and Duffy type of the red cells does not matter, as these antigens will be destroyed by the action of the enzyme).
The plasma is split into three aliquots, and each is then adsorbed by the R1R1, R2R2 or rr red cells. These are incubated at 37oC (usually, but, if a "cold" auto-antibody, or a mixed "cold"/warm auto-antibody is suspected, 4oC) for about 20 minutes. The mixture is then centrifuged, and the plasma, if initially in R1R1 red cells, transferred to another aliquot of (packed) R1R1 red cells (if R2R2 then to the next aliquot of R2R2 cells, and, if initially rr red cells, to the next aliquot of rr red cells). The incubation is repeated, as is the centrifugation and the transfer to the next aliquot of red cells.
Eventually, the uto-antibody will be adsorbed out, and then all three (individual) plasma samples are panelled against your normal antibody identification panel. Any alloantibody will then become obvious (remembering, of course, that, if the alloantibody is, say, an anti-Jka, those cells used to adsorb the auto-antibody that are also Jk(a+b-) will have taken out the anti-Jka, but that those that are Jk(a-b+) will not have done so).
Rarely, enzyme-treated red cells will not adsorb out the auto-antibody, in which case (unfortunately) you have to start all over again with non-enzyme-treated red cells. This is a complete pain, but, nonetheless, vital.
Of course, either way, you will miss an antibody such as the anti-Lan I spoke about earlier, because the chances are that all three red cells will be Lan Positive (but you can't have everything.
I hope this helps.
By the way, the correct term is adsorption, rather than absorption. If you see water vapour condense onto a window (fogging it up), this is water vapour adsorbing onto the glass surface. If you see water going into a sponge, the water is being absorbed into the sponge. One is on the surface, whilst the other is in something. The antigens on a red cell are on the surface, rather than in the red cell cytosol, and so the antibody is adsorbed onto the surface of the red cell, rather than absorbed into the red cell.
See also References (at the top of the page), choose Document Library from the drop down list, then Educational Material, go to page 2 and choose the PowerPoint lecture entitled "Laboratory Investigation od Autoimmune Haemolytic Anaemia".
:):)

It was  Polyclonal material, I didnt detect the anti C or (G) on incoming inspection but did elute it off R2R2 Cells along with an Anti D.  These days i stick to monoclonals.

You should probably get used to the DTT procedure.  Hemo-BioScience offers the reagent in small aliquots that can be used easily without bothering with trying to manufacture the stuff.  There are several threads on this already on this site.  Do a search and see the discussions.   I had posted our procedure in one - let me know if it is not accessible now and I can send it to you.
The cord panel method would be nice if you are doing a lot of pts, and at least the cells will last a while.  DDT treated cells will not last long at all.