Melanie

Hi. The question of the age. The institution I currently work at does not believe in the use of prewarm technique. We use gel as our screening method and if pos we work up the antibody in peg. If it then appears to be a CAA or something of that nature, then we do weird things like omit I.S., and repeat, if still pos, then we try 30 min no enhancement screen without IS. If not negative we use REST for untransfused people and redo screen, or differential absorption for patients who have been transfused. If we name it like Anti-M, Anti-P etc., and it is a prenatal sample, then we first decide if the antibody is IgM or IgG or both and use DTT treated serum. The antibody must react with a heterozygote cell in order for us to consider a titer and it must be clinically significant (IgG). I hope this helps you.

We give group O to all our neonates. It simplifies collection and processing and making of aliquots. It is also safer, since most transfusions are based on one type. We use leukoreduced CPD irradiated units.

So, Now I had already DTT treated some cells the day before and thought, what the heck, I'll give it a try,,, can't hurt? The three DTT treated cells were negative! AHHHHHHHHHHHH HAAAAAAAAA! What is destroyed by DTT? You guessed it..... the K system! I then typed the patient for Kpa, Kpb, Jsa, Jsb and k. The patient was Kpb neg..... Who would have thought this???? Not me. To confirm this, I thawed out some Kpb neg cells and ran them in Liss and Peg and ruled out all other common allo's . :mrgreen:

ok....This one was a real challenge to my brain. The patient was post surgical, and anemic. New eluate showed ony Anti-Fya using saline-IgG. Anti-c, Anti-Fya with Peg enhancement. So, not a WAA reacting in serum? Ran a titer of R1R1 S,K,Fya neg cell and it was 128....Probably and HTLA. Since R1R1, S, K, Fya neg cells that are neg for various HTLA's are not that available, I had to absorb and elute to rule out, I used "untreated" absorption cells and prepared eluates off the first passes. I then tested the eluates with selected cells to try and rule out. The eluates run with saline-IgG showed only Anti-c, E, Fya. When tested with Peg enhancement, I also found a Anti-S. +++++++++++++++++++++++++++++++++++++++++++++ So we now know the patient had Anti-E, c, S, Fya for sure and all other common allos except K are ruled out. So, in a pinch I sent out units that were Antigen neg "least incompatible". Now to prove what HTLA it is........ Any thoughts on that?

I hate left overs! Remember when you mother would drag yesterdays dinner and reheat it for today!!!!! and all you wanted was to start fresh, well here was one of those antibody ID left overs........ Patient recently transfused with six units of red cells in the past month. Patient requires additional units and now the antibody screen is pos, dat is pos and hospital is questioning a WAA. Patient is an O pos. Probable R1R1. DAT is micro with AhG, Micro with IgG and negative with C3b,C3d. Liss panel is set up and some cells react 3+ at imm spin and some are neg. all but one cell reacts at 37'C with different strengths from wk+ to 3+. All cells react at IgG from micro to 4+. Ficin treated cells (demonstrates a perfect little c at 37'C reacting 3+) at IgG all cells react. The previous tech had performed a differencial absorption with papain treated cells and left it for me to test. All cells were positive, some reacted at IS, some at 37'C and all at IgG. =============================================== So, now it becomes a STAT! and you are no further along then the tech was last shift, So, how would you proceed? ************************************************************* And remember, suicide or calling in sick are not options that can be entertained! ============================================== ok,, who said go home and hand it off? (only kidding) Next: since the patient appears to be an R1R1 with clean typings, I gave antigen typing a try. Some typing were weaker than controls and some were only micro. So, basically you might conclude that the patient was S, Fya neg. ++++++++++++++++++++++++++++++++++++++++++ Next I tried R1R1, S(-), Fya(-) cells and they all were neg at IS, neg at 37'C and wk+ to 1+ at IgG. +++++++++++++++++++++++++++++++++++++++++++++++ going back to the original panel you could then see that the IS reactions were Anti-E, and maybe anti-S. The Anti-K was suspicious and in ficin you know you had at least a Anti-c. But, we still have no negative cells and can not rule anything out! Eluate had been run against the origianl panel and reacted from micro to 3+ and then it was run against two R1R1 cells and two rr cells and with the R1R1 cells it was micro and it was 2+ with the rr cells. and there was no more left for me to test, So I busted up the two clots and made more. @@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@ What cells would you select to run with the new eluate? What serum testing would you do to try and rule out ? I will fill you in after some replies, thanks I hope you like these things.........Melanie

ok....i finally figured out how to get back to this site. Patient received the two compatible units with a positive outcome. The additional antibody was determined to be an HTLA. It was detroyed in ficin, and still reacted with DTT treated cells, but because it did not react with most cells tested, we could not determine which one it was and left it as "unidentified"

Patient, 57 year old male presenting with SOB due to chronic anemia. Crit of 21% Last transfusion was in 2000 and he received six units for a GI bleed. History of a non specific cold auto- agglutinin in 1999 and 2000. "don'Tcheat......Try and decide how you would proceed! Current reactions: Front types as an A pos (R1R1), Back types as a group O. Gel - all cells are 4+ and auto control is negative, Liss panel demonstrates some reativity at IS, all cells are 3+ at 37'C and 3-4+ at IgG. Peg---All cells are 4+ and auto control is neg Ficin treated cells are 3+ at 37'C and 4+ at IgG, Auto control is 2+ at 37'C and 1+ at IgG DAT.....AHg (neg), IgG (neg) C3b, C3d(neg) Cold panel......(three O cells, Cord, I neg adult, Auto and A1, A2, B cells) All cells except auto are 3+ at R.T., 4+ at 18'C and 1-2+ at 4'C auto is neg until 4'C then it reacts only 1+ ok............put your thinking caps on........what would you do next??????? @@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@ Well, I tried VEL neg, Bombay, Para-bombay and they all reacted !!!!!! REST absorbtion was a waste of time..... it weakened it by about one degree. I phenotyped the patient and ran matching cells, they all reacted the same. I then took and performed a differential absorption to rule out all "common" allo-antibodies. It took five passes, four at 37'C and one at 4'C. There were no commom allo antibodies present. ________________________________________ok!!!!! So we know it must be a high freq, but which ones would like IS,37; and IgG? and remember the patient has not been transfused in four years? Ok...throw out some ideas......Please respond and I'll tell you what it is and how I finally figured it out!

Sample sent on 67 year old male multiply transfused in october of 2003 in renal failure following a transplant who needs dialysis and transfusion ASAP. Patient has a history of Anti-E, K, Jka. A panel of cells was run that was negative for E, K, Jka and about 75% reacted from micro to 3+ using Liss and peg enhancement techniques. The auto control was 1+ in both, but the DAT was only micro using Polyspecific and IgG coombs. It was negative with Anti-C3b,C3d. Eluate did not react with any cells using both saline and peg enhancement techniques. Patient was fully phenotyped and found to be R1r, M-N+, S-s+, K-, Fy(a-b+), Jk(a-b+), Le(a-b+) The antibody that was reacting 3+ appeared to be Anti-S and was able to ruleout Fya, Lea and M. I xmatched 8 units that were E, S, K, Jka negative and 50% reacted with Liss and 80% in peg from micro to 1+. I then ran a cold panel and titered one of the incompatible units. The antibody titered out to 128 and there was also a non-specific cold auto-agglutinin at 18'C and 4'C that reacted 2-4+. I have two compatible units and all common allo-antibodies ruled out....what next? any suggestions are welcome..... Thanks

panel cells can sometimes be useful. Check extended typings before putting panel in use and hold and freeze the good ones.

titer a phenotypically matched cell, see if it acts like a HTLA. If it does and you are unable to rule-out, then you should do absorptions and elutions. The use of DTT and enzyme techniques can help you decide what raw antiseras to use to test patient cells and what cells you wish to thaw for additional testing. Sometimes you just get lucky....... We use a flowchart based on reactions with different enhancement techniques and try and follow it as best as we can. But, as we all know, the antibodies do not read the books!!!!! :| :mrgreen: