Hemolysis mystery

[QCDan]

Here is a good one we have been working on...

Patient with MDS (myelodysplastic syndrom) presented to the clinic needing a transfusion. Patient received two units of Leukorduced RBC's without complications(Hgb increased form 6.9 to 7.5). O pos, antibody screen negative (2 cell solid Phase screen galileo), electronic crossmatch performed. Patient called the clinic and came back in the following day with hemoglobinurea, blood in the urine, samples were drawn and found to be hemolysed, LDH extremly elevated, bilirubin elevated, slight lower back pain. Sounds like a classic transfusion reaction at this point. Pre- and Post transfusion samples were evaluated, testing performed included (ABORh, ABSC (gel), DC(poly)), all results remaind unchanged, O pos, negative screen, negative DC (poly). The segements of the two units that were transfused were retrived and crossmatched through gel and one of them had a 2+ incompatible result. We were at this point thinking a Low incident antigen might be causing the problem. Antibody ID was performed, samples were sent (pre and post transfusion) to the ARC reference lab. All cells had a negative reaction.

On her next go around, we did a serologic crossmatch for the two units of LRBC's she was about to get in the clinic. Antibody screen was negative in gel and solid phase. Two gel compatible O Pos LRBC's were transfused(Hgb increased from 6.7 to 7.3). Patient had no complications during the transfusion so we thought we were out of the woods. Fast Forward to the next day when she came back in with the exact same symptoms of a hemolytic transfusion reaction. The transfusion reaction work-up had the same results as it did before. Our next step is to give phenotypically similar blood for transfusion, and get HLA matched platelets in for this patient. Further down the road, since this patient will now be chronically transfused due to her MDS, we might even get one of her HLA platelet donors donate some blood for her if the phenotypically matched blood still causes her to hemolyse the transfused RBC's.

Has anyone out there ever run into something like this? It is really starting to frustrate us not being able to pin-point the cause of her hemolysis.

Thanks and I'll keep updating as we find new results.

[Yanxia]

This case is so interesting and mysterious.

I think maybe it is because some antibody in the cold circumstance will join the antigen, when in the warm circumstance ,will active comlements and hemolyze the cells, this kind of antibody will give negative screening and only C3 positive DC(this is not fit, because in this case the poly DC is neg) we will test this kind of antibody use Donath-Landsteiner test.

[Malcolm Needs ☆]

I have heard of certain Rh antibodies, in particular anti-e and anti-E, that have not been detectable in the plasma, but which must have been present, because when e or E negative blood was given, there was no delayed haemolytic transfusion reaction, but when they were given, there was always a delayed haemolytic transfusion reaction. The first time I headr about this was in a lecture given by Malcolm Beck, but I have also heard of it in a case study given by Bill Chaffe.

[tricore]

Had a patient just like this (myelodysplastic syndrome). He came in as an outpatient, negative screen, received 2 RBC, went home, chills, red urine , etc. Worked up every which way and found :(nothing.Got a new specimen several days later, found anti-K1. :cries:Continued to see this patient, antibody screen was never positive again.

Also saw an anti-c detectible only by papain and liss.

Another patient had a hemolytic reaction, found nothing other than the anti-Jkb we knew she had. About a month later a tech wentto file cbbinet and found an old workup with anti-S that had never been entered into the computer. Never had the anti-S appear on a workup again.:mad:

Did you try to ID the antibody from the incompatible crossmatch by doing an eluate on the donor cells and testing against common and rare cells?

[QCDan]

We were not able to do an elution on the Donor Cells, not enough sample left in the segment that we had. It does sound pretty similar to your transfusion reaction workup. We are still getting negative screens on this patient (3rd one now), and we have not transfused her further to this point.

I am aware of some research papers that found patients that hemolyse due to Rh, Kidd and Duffy Aby's but have a negative ABSC throughout their workup's. We are still awaiting results from the next transfusion with Phenotypically similar blood.

[John Eggington]

Are there any results for monospecific DAT tests, for example, with anti-IgA? If the patient had only an IgA class allo, or auto, antibody a 'regular' IAT, or 'poly' DAT, would not detect it (rare, but not impossible. MDS patients can throw up some 'unusual' stuff!).

Was an elution perormed on any of the patient samples? The DAT may be neagtive, but an elution may still contain detectable antibody. Although, in this instance, the DAT may be negative because all (or most) of the cells that had been coated with any antibody had been destroyed prior to testing (the samples are lysed, after all). I've also found that 'gel' DATs are good when all red cells are coated with antibody, but may not detect a 'minor' population of (even heavily coated) red cells, that may be found in a transfusion-reaction-type investigation (particularly if a lot of the coated cells have already been destroyed). A tube DAT, checked by lens/microscope, might pick up a positive 'minor' population better than a 'gel' card.

[clmergen]

There was a patient at one hospital I worked at that had a similiar reaction that we could never clearly identified. It was suspected that she possibly had an undetectable anti-c (i think deducted thru ag typing). We ended up giving her c- washed red cells and she did well.

[Eagle Eye]

I would run DAT by gel...prepare an eluate and run an eluate be gel again(we have seen tube Neg DAT and positive Eluate). I would also run extended panel by gel with low frequency antigen....eg. anti-Kpa

[QCDan]

We ran an extended panel on this patient initially looking for Kpa, He, Jsb etc. and it all came up negative, as did the gel DAT. We did not perform an elution on this patient and neither did the reference lab when we sent the sample off.

Thanks so much for all the great suggestions so far. We still have not transfused her for a third time...might be sometime this week.

[AMcCord]

We had a patient with anti-E that was detectable only when we did a ficin treated screen or panel. She also had another antibody we could see (can't remember what, though). She would react by AHG with about 1 in 3 or 1 in 4 donors which were negative for the antigen which corresponded to her other antibody, but her screens and panels were always negative. When I finally thought to try a ficin panel, I picked up a big, fat anti-E. The patient always had issues with transfusions, though thankfully not acute hemolysis, so whether or not the anti-E mattered in her case...I don't know.

[Dr. Pepper]

Those hemoglobin rises are not very impressive bumps for a non-bleeding patient - how long after the transfusions were they taken?

You have used solid phase and gel with no success - how about some other techniques? Ficin as suggested above? I also remember hearing about a case like yours where the patient was having reactions but they couldn't see an antibody until they sent it to a lab that did polybrene testing. Maybe polybrene could help you out. It probably won't help, but it can't hurt to try LISS and PEG as well. You might try a double incubation technique (thank you Peter Issitt!): set up your LISS or PEG tube test, incubate the full 30 minutes mixing the tubes a few times during the incubation, then spin and remove all the supernatant. Add a drop of saline, 2 drops serum, 2 drops enhancement medium, mix and repeat the incubation and proceed to AHG. Your cells get exposed to twice as much antibody. (I have not tried this with PEG but, assuming you can get your cells to resuspend after spinning down the first mixture, I don't see why it wouldn't work.)

I'm also intrigued by the one clearly incompatible crossmatch from the first round of transfusions. It won't explain the second bout of hemolysis, but suggests you might have 2 problems: an antibody to a low incidence antigen and a second stealthy one that you are having trouble detecting. If you can get more cells you could try further testing with that donor and at least do a DAT to eliminate the possibility of the incompatible crossmatch as being a serological red herring.

My other thought with the negative DATs is that you're doing the DATs too late, after all the cells have been lysed. Perhaps if she's admitted after the transfusions you could do serial DATs in hopes of finding some coated cells before they get zapped.

Very interesting, good luck and keep us posted.

[QCDan]

The ficin panel, PEG and LISS enhanced cells were all negative. We did an elution today on one of her samples but I left before it was completed so I will update those findings once I get them.

Some of my fellow blood bankers are thinking in the direction of an HLA incompatibility that could be causing the problem but the jury is still out on that. She has a PRA of99% and a list of HLA antibodies a mile long.

Thanks for the suggestions and input.

[NedB]

Have you run an IgA level on the Patient? The Patient may be showing IgA sensitivity, along with the stealth antibody.

[Malcolm Needs ☆]

The ficin panel, PEG and LISS enhanced cells were all negative. We did an elution today on one of her samples but I left before it was completed so I will update those findings once I get them.

Some of my fellow blood bankers are thinking in the direction of an HLA incompatibility that could be causing the problem but the jury is still out on that. She has a PRA of99% and a list of HLA antibodies a mile long.

Thanks for the suggestions and input.

This is quite interesting. There is little doubt that, although extremely rare, HLA antibodies can be associatedwith red cell destruction.

If you go to the top of the page and click on "Library", and then choose "Educational Material", and then go down to "Blood Transfusion in Haemoglobinopathies" (or a title very like that), you will see a PowerPoint lecture and accompanying Word document written by my Consultant, Dr. Nay Win, that explains this in a few slides starting at number 70.

Although here he is talking about such antibodies causing hyperhaemolysis in sickle cell patients, the same would apply to other patients, and I know that he wrote a paper on a fatal case of hyperhaemolysis in a patient with MDS (I may even have been a co-author myself - I can't remember, but I have been a co-author on a very few of his papers on hyperhaemolysis!).

It may well be worth trying HLA-matched, washed red cells and seeing how the patient tolerates these, and, maybe, covering with IVIG and methylprednisolone.

:idea::idea:

[rravkin@aol.com]

QCDan,

Has any consideration been given to a mechanism of RBC destruction outside of immune response where by this patient's condition may predispose for such post transfusion response. As Phil has mentioned in his amazing post "Those hemoglobin rises are not very impressive bumps..." and they are not!

Edited December 29, 2010 by rravkin@aol.com

[Colin Barber]

Dear QCDan,

I have a few questions and a suggestion for another line of thought:

1) Is the ARC part of your Transfusion service ? were their findings like yours no aparent red cell antibodies in the pre & post transfusion samples.

2) If the ARC is part of your donor blood collection service ? do they have any cells from donor testing with which they can reapeat the crossmatch on any units given and perhaps to a DAT on the unit that was incompatible (was that with pre & post transfusion plasma).

3) Did the post transfusion Hb level fall below the pre transfusion level.

Point 3 leads to my alternative line of thought - If the post transfusion Hb fell below the pre transfusion Hb then is this a case of transfusion induced hyperhaemolysis. Whilst this is normally seen in Sickle Cell or Thalassaemic patients it has been described in non-haemoglobinopathy patients as well.

I have seen several cases of transfusion induced hyperhaemolysis in our sickle patients, but I did have a haemophilia patient who I am convinced also had transfusion induced hyperhaemolysis - his serology was negative and his Hb crashed following transfusions with all other symptons of an acute haemolytic episode. He has not required any red cell transfusion recently so I have not had a chance to explore this theory with him further.

Good luck with your MDS patient.

Colin

P.S. I should have read all the posts on this before I posted my thoughts - I see Malcolm has also mentioned Hyperhaemolysis as well.

Edited December 29, 2010 by Colin Barber

[Malcolm Needs ☆]

I should, perhaps, also warn that hyperhaemolysis can be fatal, and can be recurrent, and so if this condition is a serious contender in this (or any other) case, further transfusion should be avoided unless the anaemia becomes life-threatening, until hyperhaemolysis has been disproved.

If transfusion is required before disproving hyperhaemolysis, or indeed, if hyperhaemolysis has been proved, I would seriously suggest that an expert in the subject is consulted first, and would also seriously suggest, as I did above, that consideration is given to covering the transfusion with IVIG and steroids.

I should alsao mention that there are some very high profiledoctors who do not believe that hyperhaemolysis syndrome exists in the first place.

:omg::omg:

[rravkin@aol.com]

QCDan,

Is the hematuria in this case caused by presense of free hemoglobin, intact red cells, or a combination of both?

[QCDan]

Here are a few answers,

-The ARC refernece lab is our contracted helper in cases where we have limited resources to get to an answer of a immunohematological problem. The had the same findings as we did concerning the ABID work-up and DAT.

-The DAT on the incompatible unit was negative.

-That particular unit was incompatible in Gel technology with the pre, and post transfusion sample on the first go- around.

-An elution was performed recently on a fresh sample and was negative (we tested for low fequency antigents also).

-In the matter of the hyperhaemolytic syndrom, I don't think that is the case here since this person did not have a decrease in Hgb until three days after the transfusion and even then, it was from 7.2 to 6.2. two days later it fell to 5.8, at which point the patient was transfused for the second time with 2 units of RBC's giving a rise in Hgb to 7.4 (pretty standard increase) but she was at this opint experiencing yet another transfusion reaction. We have not transfused her since then and her Hgb is currently at 7.9

-As far as I know there was hemoglobin in her urine, I'd have to check back though some of her results to find out if intact RBC's were also present.

Thats all I have so far. Her Platelet count is also dropping and we are obtaining HLA matched Plt's for this patient but she has not received them yet.

Thanks

[Colin Barber]

QCdan,

I am not so sure I would dismiss hyperhaemolysis so readily - in your original post describe an agressive haemolytic process which from your description sounds like it was intravascular - back pain, Hb in the the urine, evidence of haemolysis in the post transfusion sample. You also said the second haemolytic reaction was the next day. There may well be an antibody to a low frequency antigen which could explain the incompatiblity found to the unit in the first transfusion reaction you describe, but as other have said this does not explain this patients continued post transfusion haemolytic episodes.

In the Sickle patients I have seen with Hyperhaemolysis they usually have allo antibodies and often an auto as well, so red cell antibodies may well be present in your patient. The Haemophilia patient who I think had hyperhaemolysis did have strong multiple HLA antibodies - which does tie in with what Malcolm was saying about Hyperhaemolysis.

Your patient is obviously going to need continued transfusion support, so lets hope you do get a definitive answer.

Colin

Edited December 30, 2010 by Colin Barber

[Yanxia]

What is hyperhemolysis?

[Colin Barber]

What is hyperhemolysis?

Shily, I am sure Malcolm will also post a reply as I know he has investigated cases of Hyperhaemolysis. I my area of London we have a number of Sickle Cell patients who are treated in our hospital. Within that population of Sickle Cell patients we have at least 4 that I know of who are not able to be transfused.

The reason for this is whenever they have been transfused once they have become sensitised they have a haemolytic transfusion reaction in which they seem to destroy not only the the transfused cells but also some of there own cells as well.

It's very frightening when it happens because as Malcolm says it's potentially fatal. As far as I know there are lots of theories but no one knows the actual mechanism involved in the haemolytic episode, it appears to be an acute onset aggressive haemolytic process triggered by transfusion. Most of the patients have complex serology usually with multiple allo antibodies, the haemolytic reaction occur when phenotyped and crossmatch compatible units are given and post transfusion reaction investigations still show that the units were apparently compatible.

I think there is a PhD for who ever works out exactly what is going on in these patients.

Colin

[Malcolm Needs ☆]

[ATTACH]447[/ATTACH][ATTACH[ATTACH]447

IF I have attached these files properly, then they may be of use.

The PowerPoint was written by Dr. Nay Win, my Consultant, who is an expert in the field of hyperhaemolysis, and originally suggested the theory of hyperactive macrophages, a theory that is rapidly gaining support throughout the world.

The particular slides that may be of interest are from 61 to about 85.

:fingerscr:fingerscr:fingerscr:fingerscr:fingerscr

Transfusion of Sickle and Thalassaemic Patients.zip

Transfusion of Sickle and Thalassaemic Patients.doc

Edited January 2, 2011 by Malcolm Needs
Can't count!

[rravkin@aol.com]

Malcolm,

I am sorry that I do not have the moment to review this article prior to asking this question; but are these presumed hyperactive macrophages located in the Spleen? It would make sence if they were given that the Spleen maintains a large population of Macrophages and the transfused RBC's go through a sequestering period in the Spleen before entering the circulation for the remainder of their circulating life.

I was speaking with a co-worker about this case today and she was telling me about a similar experience with a patient whereby they desided to transfuse 100ml aliquots of PRBC's instead of a whole unit at once. Upon doing so there was minimal to no RBC destruction but there was the expected minimal increase in HgB after the first couple of aliquots. Is there any practice like this given in the litererature or that you know of when working with a case like this?

Edited January 3, 2011 by rravkin@aol.com

[Malcolm Needs ☆]

Malcolm,

I am sorry that I do not have the moment to review this article prior to asking this question; but are these presumed hyperactive macrophages located in the Spleen? It would make sence if they were given that the Spleen maintains a large population of Macrophages and the transfused RBC's go through a sequestering period in the Spleen before entering the circulation for the remainder of their circulating life.

I was speaking with a co-worker about this case today and she was telling me about a similar experience with a patient whereby they desided to transfuse 100ml aliquots of PRBC's instead of a whole unit at once. Upon doing so there was minimal to no RBC destruction but there was the expected minimal increase in HgB after the first couple of aliquots. Is there any practice like this given in the litererature or that you know of when working with a case like this?

Hi,

Just to make certain that my reply is correct, I think it best if I talk to Nay Win before I answer your specific points. I should be seeing him on Tuesday.

Malcolm

:whisper::whisper: