auto little e ???

[phoenix_1619]

Hello to every body.. Please be patient while reading this long thread... but I really need your help.

we have a 4 years old patient with sickle cell anemia and multiple sequestration for seplenectomy

her previous history in the referring hospital is that she is A RhD pos ( cells are reacting only 1+ with anti D) by tube method and she has been transfused 12 times by A RhD Negative and she has no phenotype , her previous ab screen was Negative all the times . The last time that they did it was the first of April and she had her transfusion with no problems.

She was admitted on the third of May with positive Ab scr ,3 cells were 1+ and AC 1+, DAT was 1+, IgG 1+, C3d Neg .Ab ID has one cell negative which match with the (e) , I could not rule out the C, K,S, Fya, JKa and e

after acid elution a panel was run and all the 11 cells were reacting 3+.

her phenotype was R0r , i.e: E,C Negative and e, c Positive , this was done by Diamed Id cards using her cells after treating with hypotonic saline to hemolyse the donor cells ( there were actually two cell populations) so our pathologist advice her doctor to wait for one month later to have a clear picture of her phenotype.

At this time we started to use the cards for most of our testing.

After she came again last week through OPD clinic we phenotype her again , the Rh phenotype was same as before R0r

she is Kell , M,N,Jka,Jkb, P1 positive , i couldn't phenotype the Ss , k & Duffys because her DAT was positive. i tried the old fashion gentle heat elution method to elute the Ab while saving the integrity of the cell but it didn't work, the DAT was still positive. her Ab scr, AC, DAT, IgG, Ab Id are all reacting 2+ by the cards , I did the auto adsorption by PEG but only twice not triple ( i have little cells) the auto adsorbed sera was reacting exactly as the little e ( one cell negative when (e) is Negative , one cell is 1+ when (e) and E are both positive "heterozygous" while all other cells are reacting 2+

Sorry for the details and now coming to my Questions:

1- our pathologist said that the reaction in the cards looks like flocculation not true agglutination and is not convinced that it is auto Ab with specificity because the patient is not hemolysing ( we are new to the gel method so could some one explain this flocculation issue to me please?)

2- What is the best way to phenotype the patient cells? If possible kindly suggest a reagent that can help with the name of its manufacturer, so we can order it for future use.

3- What is the best choice of phenotype that I have to give her if transfusion is decided?

Thanks in advance to all the helpful ideas:confused:

Eman

Edited June 7, 2009 by phoenix_1619
add some thing

[Yanxia]

Chloroquine phosphate can eluate the antibodies and keep the cell's integrity.

[Malcolm Needs ☆]

Hi pheonix_1619,

I would agree with shily that chloroquine diphosphate should help you to resolve this child's phenotype, but you must be aware, if you do decide to use it, that it does substantially affect some antigens, which are rendered very weak indeed. These include Mt(a) (a very rare antigen within the MNS Blood Group System, so probably not a worry here), Fy(, Lu( and Yt(a) (again, probably not a worry in this case, but more to the point antigens within the Rh Blood Group System can also demonstrate weakening (althoough not to the same extent), together with antigens of the Dombrock, Knops and JMH Blood Group Systems (once again, probably not a worry in this case). If it is available to you, at your local Reference Laboratory, a much better approach would be to get the patient genotyped, but I realise that this method is not available to everyone.

Not having seen the reactions (obviously!) I am going to stick my neck out here, but I'm not entirely convinced that your Pathologist is correct in saying that the reactions are are result of flocculation. Were this to be the case, one would expect such reactions with all red cells tested, and you are not getting such reactions. Indeed, the reactions you are getting seem, on the face of it, to be an auto-anti-e.

The fact that the patient has a positive direct antiglobulin test supports the theory of an auto-antibody. It must be remembered that not all auto-antibodies cause clinical haemolysis. It could be that her own red cells are not remaining in her circulation for as long as would be expected (probable anyway, as she is a sickle patient), but that there is a compensated anaemia, which masks the haemolytic process.

I doubt very much if the auto-antibody is actually anti-e, but is much more likely to be an auto-anti-e-like antibody that is mimicking an anti-e. You can test for this quite easily by adsorbing the antibody with red cells that are e+ and red cells that are e-. The antibody will, obviously, be adsorbed out using the e+ red cells, but if you persist, you will probably find that the e- red cells will also adsorb out the apparent anti-e eventually. You have to be very careful here, in that you have to be able to demonstrate that the antibody has not been diluted out with repeated adsorptions. Try adsorbing out another patient-derived antibody of a different specificity (a genuine specificity - say an anti-K) of similar strength of reaction, and adsorb this in parallel with (in the example given) a K- red cell. If this antibody is still detectable after the same number of adsorptions, then it is less likely that the apparent anti-e has been diluted out.

Lastly, whilst K+ is not unknown in sickle cell patients, it is very rare, and so I would be very inclined to repeat this typing, just to make sure.

This is by no means a definitive answer, and I suspect that other people will have different, better, suggestions to make, but I hope it will be of some help to you.

Edited June 8, 2009 by Malcolm Needs
Spelling - as usual!

[David Saikin]

Many autoabs will have an anti-e-like specificity if you dilute them enough. Malcolm gives good advice. I would not do all that work though. Even if you give e= cells, the ab will probably coat them and result in decreased rbc survival. A true autoab will treat the transfused cells just like the pt's, i.e., will cause decreased survival but not overt hemolysis.

[Malcolm Needs ☆]

I agre entirely with what you say David and, candidly, NEITHER WOULD mALCOLM normally do all that work either!

I would, however, do at least some of it in this case, just to reassure the Pathologist that what I have said may be true, so that I wouldn't have the same problem in the future. I probably haven't worded that very well, but I know what I mean!

Edited June 8, 2009 by Malcolm Needs

[galvania]

I would also advise you to give her Fya- blood, as she has a very good chance of being Fya-b-. Poor little thing. Only 4 and already having to go through all that! Incidentally, I would love to know what your pathologist means by 'flocculation' in gel. I've been working with gel for nearly 20 years and it's the first time I've ever heard the term in this context.....

[BSIPHERD]

There is a method in the AABB Technical Manual (p. 897) to separate transfused red cells from autologous red cells in patients with hemoglobin S disease (uses hypotonic saline).

Also, the little e antigen is pretty complex expecially in blacks. Sometimes what appears to be autoanti-e, particularly in sickle cell patients, turns out to be some form of alloanti-e. I know they've worked at lot with these at the Red Cross in Philadelphia. I can't really give you more information, but I know it's out there.

[Malcolm Needs ☆]

There is a method in the AABB Technical Manual (p. 897) to separate transfused red cells from autologous red cells in patients with hemoglobin S disease (uses hypotonic saline).

Also, the little e antigen is pretty complex expecially in blacks. Sometimes what appears to be autoanti-e, particularly in sickle cell patients, turns out to be some form of alloanti-e. I know they've worked at lot with these at the Red Cross in Philadelphia. I can't really give you more information, but I know it's out there.

What BSIPHERD, 15631 says about the e antigen amongst the Black population is absolutely true and, more to the point, genotyping will not necessarily help, as the whole thing is such a mess.

Heterogeneous molecular background of the weak C, VS+, hrB–, HrB– phenotype in black persons (p 495-504)

Bach-Nga Pham, Thierry Peyrard, Genevieve Juszczak, Isabelle Dubeaux, Dominique Gien, Antoine Blancher, Jean-Pierre Cartron, Philippe Rouger, Pierre-Yves Le Pennec

Transfusion 2009.

Edited June 8, 2009 by Malcolm Needs

[phoenix_1619]

Great thanks to you all for the replies

i got a good ideas and i'll do some of it today.

i called the product specialist in Diamed and he is coming to help me in resolving this flocculation issue !!

my pathologist used to work in a hospital that use the gel long time ago but this is her fourth year here in this hospital that use tube method. she said that the positive is showing a clear band .

our patient is not black and i'll send her sample for genotyping in her next visit if DAT remain positive.

thanks again and see you later

[Malcolm Needs ☆]

The point is phenonix_1619, your patient may not actually be physically black, but with her probably Rh phenotype of Ro, and her sickle cell disease, she may still have multiple genes from the Black population, so she may still have an atypical e antigen.

[Townsend]

I'm the TS Manager at a Children's Hospital with a large sickle cell population. For some time now, we have had a phenotypically similar blood program in place, and have had quite a few patients that fit your exact description. Here are my thoughts:

- Have you considered sending this patient to a reference lab for Molecular Testing to obtain a true phenotype? We send samples to our local ARC Reference Lab for a phenotype upon initial diagnosis (to help avoid these situations, where after multiple txns, it's hard to obtain the patient's phenotype). Anything requiring molecular testing is sent on to ARC, Penn-Jersey Ref Lab. We have a lot of patients who have made Warm Autoantibodies that mimic anti-e.......... the question is, is it a true auto-e, or does the patient have a varient e antigen? In that case, the e antigen typing would be positive, but the patient forms an allo-anti-e, against the "traditional" e antigen. Much more common in our sickle cell patients than you'd think. In the meantime, as recommended by our Ref Lab folks, I would try to honor the e and give e neg units until you know whether or not it is allo or auto in nature.

-So, based on your results, we would try to obtain blood that is negative for C, e, Fya, S negative which is a "likely phenotype" for these sickle patients. If the e turns out truely to be auto, you may be able to drop the e neg requirement and give units based on the true phenotype. I would suggest matching as much of the phenotype as you can based on blood availability.

-As for the flocculation, I too have not heard that particular term. We don't currently do typing in gel, but use it for ABID/AS/DAT/AHG XM.

Hope this helps!

Stephanie Townsend, MT(ASCP)SBB

[janet]

Our sickle cell patient had (what appeared to be) an auto-anti-e also (we too use gel - I don't think we questioned 'funny' looking reactions) .....started out as a weak unidentified, progressed to a perfect looking e which about 6 months latter went away. There is literature that states 35% (I think??) of sickle cell patients will develop auto-antibodies. We felt best giving him e negative (therefore compatible units) while the antibody was there ... now with a negative antibody screen we have gone back to giving him E-,C-K- (matching his phenotype).

Just thought I'd add my experience - sorry, not an answer to your questions :0)

[Malcolm Needs ☆]

Our sickle cell patient had (what appeared to be) an auto-anti-e also (we too use gel - I don't think we questioned 'funny' looking reactions) .....started out as a weak unidentified, progressed to a perfect looking e which about 6 months latter went away. There is literature that states 35% (I think??) of sickle cell patients will develop auto-antibodies. We felt best giving him e negative (therefore compatible units) while the antibody was there ... now with a negative antibody screen we have gone back to giving him E-,C-K- (matching his phenotype).

Just thought I'd add my experience - sorry, not an answer to your questions :0)

I am very sorry to say this Janet, but I must disagree with this approach, much as it seems to have worked in this case.:o:o

As I said in an earlier post, although these antibodies may appear to be genuine specificities, and often they appear to be auto-anti-e, they are, in fact, mimicking antibodies. In this case, you were giving your patient E+ blood transfusions (presumably R2R2) These red cells, however, would exhibit this e-like antigen, but to a lesser extent than would an e+ unit.

Almost certainly, therefore, you would be giving a Black sickle cell patient transfusions from a White donor (as R2R2 is less common in the Black population [0.2%] than the White population [2.3%]), and the more blood you transfuse, the more likely a recipient is to produce atypical alloantibodies).

In this case, therefore, you would be exposing your patient to the E antigen on a number of occasions, giving their immune system every chance to produce alloanti-E. In addition though, you would almost certainly be exposing them to the Jk( antigen on multiple occasions (this antigen occurs much more in the White population [73.7%] than in the Black [48.9%]) to mention but one antigen that differs in frequency between the two ethnic groups.

Sickle cell patients, like thalassaemic patients, tend to make no atypical antibodies or make every antibody under the Sun, and your may have been in the former group, but it is risk I wouldn't be inclined to take. In a study published by Vichinsky et al in 1990 (Alloimmunization in sickle cell anaemia and transfusion of racially unmatched blood. N Engl J Med; 322: 1617-1622) 50% of patients who had been given racially unmatched blood transfusions developed alloantibodies after 100 or more transfusions.

Given that all individuals (apart from those with Rh deletions) will exhibit the e-like antigen on their red cells, unless the (genuine) e+ transfused red cells are destroyed in the recipient at such a rate as to drastically increase the need for transfusion (e.g. by drastically lessening the time between which transfusions are required) it would be far better for the patient, in the long run, to continue to transfuse e+, E- red cells in most situations.

Having said all of that, I doubt very much if 100% of the contributers to this website would totally agree with me!:disbelief

[Brenda K Hutson]

Just a few thoughts from reading your posting:

1. Just because a patient has an autoanitbody, even one with a specificity like anti-e, does not mean

they are going to hemolyze. In my 26 years and 6 Institutions, I have seen that the majority of

autoantibodies (with or without specificity) are not hemolytic.

2. I think the standard of practice in "most" Institutions these days, is that you do not transfuse an

auto-e patient with e-negative blood, unless they are hemolyzing.

3. There are 2 reasons for #2: First of all, you don't want to waste precious e- blood for an

autoantibody; you need to save it for the allo anti-e patients. And second, keep in mind that when

you are giving e-negative blood, conversly, you are giving E+ blood (and this patient is E-). If you

stimulate them to make the allo-E, you now have the dilemna of a patient with an auto-e and allo-E

which could become problematic, depending on what is going on with the patient.

4. I have seen some pretty ugly work-ups on Sickle Cell patients who have made variant anti-e.

5. I don't have the Instructions in front of me, but you can purchase EGA or Chloroquin to strip antibody

from DAT+ RBCs; this will allow you to phenotype (though if DAT is strong, this may not be

successful)

Just my thoughts,

Brenda Hutson, CLS(ASCP)SBB

[Malcolm Needs ☆]

Agree 100% Brenda.

[Brenda K Hutson]

Ooops, forgot one more point in my response. The Standard Protocol for transfusion of Sickle Cell Patients is (and this is assuming you have determined the complete phenotype of course):

Give Rh and Kell System matched if no Alloantibodies ID. When/if patient makes Alloantibodies, give total phenotypically matched. I think you said the only thing found is an "Auto-e;" correct?

Brenda Hutson

[irshadaad]

1.in diamed gel cards you cant do phenotype if DAT is positive,you cant rule out any anigen which needs coob's mthod

2.since your pt is transfused many times you cannot rule out the real phenotype of the pt.

3.if u can use DTT .method for removal of antibodies from the cells of the pt to make DAT neg.the proced for any type of grouping

4 try elution and adsorption ....but not with the cells of the pt,

5.best will be rh negative blood with C - E- e-

6.PLS look for other antibodies from other specificities...have you done A1 lectine,

7 what about penal with enzyme ,rt and 4c.

8.your reaction in card is doule pop so sugests reaction with transfused cells aswell

9 try tube method with monoclonal anti sera eg SERACLONE/diamed too

10.its because in the first time may be the phenotype was not taken into consideration so afterwards heterozygous blood created anti body

hope it will help u

[irshadaad]

Just a few thoughts from reading your posting:

1.the practice in transfusion service is to provide a compatible blood and we have to try it to our best to provide blood which is negative for any antigen aginst which there is anti body whether allo or auto...its not about waisting e - blood its about not to allow antibody to attach to the cell.....especialy in sickle cell pt.viz renders finaly the cells to a short life span in vivo.....though brisk heamolysis takes place in case of compliment binding antibodies but...here too if we give e+ve blood it will subject to delayed transfusion reaction type ...why we have to do it delebratly.....when as per mollison any blood is considered incompatible which is hindered by antibodies in the circulation

[Malcolm Needs ☆]

Just a few thoughts from reading your posting:

1.the practice in transfusion service is to provide a compatible blood and we have to try it to our best to provide blood which is negative for any antigen aginst which there is anti body whether allo or auto...its not about waisting e - blood its about not to allow antibody to attach to the cell.....especialy in sickle cell pt.viz renders finaly the cells to a short life span in vivo.....though brisk heamolysis takes place in case of compliment binding antibodies but...here too if we give e+ve blood it will subject to delayed transfusion reaction type ...why we have to do it delebratly.....when as per mollison any blood is considered incompatible which is hindered by antibodies in the circulation

Nonsense.

See my replies to your other posts on the subject of auto-antibodies (and read Petz and Garratty on the subject; you may learn something).

:mad::mad::mad:

[eric1980]

I like controversies. Those who are open to controversies are usually the people who will think of more possbilities and usually will make the world a better place. Those who dismisses controversies without finding out more are usually people who are SOP fanboys/fangirls and will only possess limited knowledge because of the need to be spoon-fed with information.</end of "huh?!" speech>

In my BB, if the patient have auto anti-e or anti-e mimick, we will transfuse e- RBCs. I had always thought this makes sense. But as what Brenda had said and what I have experienced, e- RBCs are indeed very rare. And to allow a patient with those antibodies to transfuse e+ RBC is really making my heart beat faster and unable to sleep at night. =S

I will ponder this tonight in bed. =S

[Malcolm Needs ☆]

I like controversies. Those who are open to controversies are usually the people who will think of more possbilities and usually will make the world a better place. Those who dismisses controversies without finding out more are usually people who are SOP fanboys/fangirls and will only possess limited knowledge because of the need to be spoon-fed with information.</end of "huh?!" speech>

In my BB, if the patient have auto anti-e or anti-e mimick, we will transfuse e- RBCs. I had always thought this makes sense. But as what Brenda had said and what I have experienced, e- RBCs are indeed very rare. And to allow a patient with those antibodies to transfuse e+ RBC is really making my heart beat faster and unable to sleep at night. =S

I will ponder this tonight in bed. =S

Hi eric1980.

For the rationale behind my post above, read my post in the thread ALL OTHER TOPICS, AUTOANTIBODY CROSSMATCH ADVICE. I'm not trying to push what I think. This quotes the AABB Manual.

Read it (it will only take two minutes at most) and then sleep soundly!

:)

Edited July 23, 2009 by Malcolm Needs
Wrote "post" as "posy"!

[eric1980]

Thanks, Malcom. I will take a look at it. ; )

[Brenda K Hutson]

Just a few thoughts from reading your posting:

1.the practice in transfusion service is to provide a compatible blood and we have to try it to our best to provide blood which is negative for any antigen aginst which there is anti body whether allo or auto...its not about waisting e - blood its about not to allow antibody to attach to the cell.....especialy in sickle cell pt.viz renders finaly the cells to a short life span in vivo.....though brisk heamolysis takes place in case of compliment binding antibodies but...here too if we give e+ve blood it will subject to delayed transfusion reaction type ...why we have to do it delebratly.....when as per mollison any blood is considered incompatible which is hindered by antibodies in the circulation

If a patient is hemolysing due to their Warm Autoantibody (whether specificity or not), then yes, I agree you would want to give e-. However, I believe I am correct in saying that it is the Standard of Practice in most places, that if they are not hemolyzing, you do not give e- for a Warm Auto-e for 2 reasons:

1. That means you are giving them E+ which for them, is an Alloantibody; you could then make things worse if they

become sensitized to this

2. It would be considered wasting precious e- blood if you give it to a patient with a Warm Auto-e who is not

hemolyzing (at least by most people/Institutions).

And one other thing to keep in mind with the Sickle Cell Patients; when they make Anti-e, it can be a Variant Anti-e, making available blood, difficult to find (we had a patient with a variant anti-e, plus multiple other antibodies; only available blood was in Africa; Hgb between 2-3; patient sent to Hospice).

I stand by my process statements (but you are certainly welcome to your own).

Thanks

Brenda Hutson, CLS(ASCP)SBB

[Brenda K Hutson]

Nonsense.

See my replies to your other posts on the subject of auto-antibodies (and read Petz and Garratty on the subject; you may learn something).

:mad::mad::mad:

Just for clarification Malcolm, to whose e-mail are you directing the "nonsense?" To mine, or to irshadaad?? It almost looks like what is in the box on the response from irshadaad, is suppose to represent "my" previous response; but it is not.

Just curious; no hard feelings.

Thanks,

Brenda Hutson, CLS(ASCP)SBB

[Malcolm Needs ☆]

Just for clarification Malcolm, to whose e-mail are you directing the "nonsense?" To mine, or to irshadaad?? It almost looks like what is in the box on the response from irshadaad, is suppose to represent "my" previous response; but it is not.

Just curious; no hard feelings.

Thanks,

Brenda Hutson, CLS(ASCP)SBB

I'm extremely sorry Brenda.

I think waht happened was that we were posting almost together, and you just won! As a result, your post appeared just above mine, and NO, it most certainly was NOT aimed at your post.