Henrique

I would most strongly advise you to send a sample, possibly even multiple samples throughout the pregnancy, to a Reference Laboratory.

As the patient is pregnant, there is the possibility that the Jk(a) antigen you are detecting is actually being expressed on the red cells of the foetus, and you are detecting it as a result of a foeto-maternal haemorrhage.  However, the Jk(a) antigen is not necessarily straight forward, as there are weakened forms of the antigen (and the Jk(b) antigen come to that) where there are amino acid substitutions remote from the site usually associated with the Jk(a) and Jk(b) antigens (280 of the mature protein).
In addition though, you have, obviously, to consider the health of the unborn baby who, even if the antibody does turn out to be a maternal auto-anti-Jka, may cause haemolytic disease of the foetus and newborn, albeit this will usually be be very mild.
I attach a PowerPoint which may, or may not help you in your decision to send a sample to your local Reference Laboratory (also tell them the ethnicity of the patient).
 
Interesting case - please keep us informed.
In Depth Lecture on The Kidd Blood Group System.pptx

A lecturer I listened to discussed MTP and stated that using Rh positive packed cells keeps the patient alive.  He said that if anti-D is built, it can be dealt with when the woman gets pregnant.  If she dies because she didn't get transfused with Rh positive packed cells, she certainly won't even have the opportunity to become pregnant.  So, there's that.

I am a little worried about this because, even if there are VERY low numbers of isoagglutinins, if there is a normal level of complement, it can still be extremely dangerous, as there is massive amplification within the complement system (see the excellent attached PowerPoint lecture, written for me by my friend Grant Webb).
The Complement System.pptx

Erythrocytes cannot be either homozygous or heterozygous (or hemizygous for that matter).  The terms homozygous, heterozygous and hemizygous should only be used when referring to genes, and, while antigens are (ultimately) derived from genes, many, such as A, B, H, I, i, Lea and Leb are not proteins, and cannot, therefore, be direct gene products (although even the "protein" antigens go through post-translational modification and so also cannot, strictly speaking, be direct gene products).  On top of this, of course, the mature erythrocyte has exuded all nuclear material.
Antigens, therefore, should only be referred to a single or double "dose".

Hi MinerJ,
We used to do something similar to you when I was in RCI at Tooting, except that we performed tube IAT strictly at 37oC, with warmed NISS for the washing process.  As far as this is concerned, I could never see any difference between having the red cells initially suspended in NISS or LISS, except, of course, that the incubation time was shorter with LISS (although we used to bring the reactants to 37oC before the initial mixing).  Again, like you, this was mainly for pregnant women.
After many experiments trying to get the CAT reaction chamber strictly up to 37oC BEFORE the addition of the reactants, and failing miserably, we abandoned the idea, as we could never quite get to 37oC.  Unfortunately, "cold reacting" antibodies can (and do) sensitise their cognate antigens extremely quickly, but are much slower to dissociate, so we were never quite sure whether the anti-M was reacting genuinely at 37oC or not.
I think you may have made a typo.  There are no enzymes involved, because of course, the M antigen is papain sensitive, by the columns have a slightly low pH, and this, together with centrifugation and the difficulty in keeping the reaction chambers at strictly 37oC all mitigate towards the possibility of a false positive at 37oC.
I hope this bit of rambling answers your question sufficiently, but, if not, do not hesitate to get back to me (or anyone else, come to that!).
Malcolm

In terms of the function of the various ABO blood types, there have been a huge number of peer-reviewed papers written on the subject (and the number has exploded with the advent of COVID19).  I would seriously defy anyone to keep up with all of these, but I would recommend reading pages 42-43 of Reid ME, Lomas-Francis C, Olsson ML.  The Blood Group Antigen FactsBook.  3rd edition, 2012. Academic Press.  ISBN: 978-0-12-415849-8.

In terms of how they evolved, it is so far back now that it is anyone's guess, but slides 28 to 32 of the attached lecture may give you some idea.
In Depth Lecture on The ABO and H Blood Group Systems.pptx

I prefer to start with more cheaper reagent.

It would be really useful if you could tell us the ethnicity and age of the patient, and his medication regime.
That having been said, I note that the antibody screen is positive, that his DAT is positive by both anti-IgG and anti-C3d, that the neat plasma contains an apparent anti-E and anti-c, but that the eluate contains an antibody that is, apparently, pan-reactive.
Very often in these cases, the apparent antibody specificity in the neat plasma is a mimicking specificity, rather than a true specificity.  In such cases, the apparent specificity in the neat plasma can be adsorbed out using red cells that are negative for the antigens of the apparent specificity; in this case R1R1.  The true specificity of the antibody could be an anti-Rh17 or anti-Rh18.
While I am not saying for a single second that the apparent specificities of anti-E and anti-c are not true specificities, it may be worth your while seeing if they can be adsorbed out using R1R1 red cells.  However, as you suspect the presence of other antibodies, this should not be attempted until you have proved otherwise.  This you can do, as you suggest, by alloadsorption of the neat plasma using two or three adsorption cell types.
In answer to your last question, with regard to adsorption of the eluate, this was certainly a method we used in the Reference Laboratories of the NHSBT in the UK.  It was usually used when the patient had a known pan-reactive autoantibody, but was requiring transfusions more frequently than previously, and/or when the expected rise in the haemoglobin concentration was not achieved.  On some occasions, we were able to detect a de novo alloantibody in the eluate that we could not detect in either the neat plasma, or the adsorbed plasma, although this was not always the case, as transfusion in and of itself can sometimes stimulate the autoantibody to become more active (see Petz LD, Garratty G.  Immune Hemolytic Anemias.  2nd edition, 2004, Churchill-Livingstone).
Good luck with sorting it out, but this is a really interesting case.  Thank you for posting it and, please, would you mind letting us know how you get on?

Before I attempt to answer your query, I must explain that I am NOT a doctor.  I am what is called in the UK, a Biomedical Scientist and, as such, am not qualified to make a diagnosis, but I am the Chief Examiner in Transfusion Science for the Institute of Biomedical Science, and used to by the Reference Laboratory Manager in the Red Cell Reference Laboratory in the National Health Service Blood and Transplant Centre in Tooting, London, so I can claim some expertise.
Although a warm auto-antibody in a person's  plasma is by no means common, it is something we use to see on a daily basis at Tooting.  To put it at its most basic, it results from your immune system producing an antibody directed against a red cell antigen expressed upon your own red cells, which could, under certain circumstances, lead to you becoming (usually mildly) anaemic.
The "autologous adsorption" bit means that the laboratory, either at your hospital, or at a Reference Centre has been able to remove the antibody from the plasma in your blood sample by using  your own red cells (thus proving beyond doubt that the antibody is indeed an auto-antibody).  They have then tested this adsorbed plasma in tests to see if there are any unusual antibodies in your plasma that are directed against antigens expressed on the red cells of other individuals; so called allo-antibodies.  They include in their report the caveat that concerning the "common blood group antigens" because it is all but impossible to test for antibodies against all the known antigens, of which there are well over 600, some of which are incredibly rare.
Most auto-antibodies have a specificity within the Rh Blood Group System, which, at present, contains 55 different antigens (but other antigens are being found on a regular basis).  Most of these auto-antibodies are directed against either the Rh antigen known as Rh17, or against that known as Rh18 (I realise these names will mean nothing to you - but bear with me).  Almost everybody in the world expresses both of these antigens on there red cells, and the actual specificity of the auto-antibody is not really of any consequence.
It is highly unusual, to say the least, for a maternal auto-antibody to cause any problems with a condition known as haemolytic disease of the foetus and new-born (or HDFN), particularly at an early stage of pregnancy.
To me, this suggests that your early miscarriages and your auto-antibody status are coincidental, rather than the auto-antibody being the cause of your early miscarriages.  Red cells are not really produced in early foetal life (indeed, there is not much in the way of blood in a foetus until about 12 weeks of gestation), so there are very few foetal red cells available to be affected by your auto-antibody.
Having said all of that, I would reiterate that I am NOT a doctor, and even if I were, it would be impossible (and stupid in the extreme) to even attempt to make a diagnosis without FULL knowledge of your case.  As such, I would suggest that you do discuss your case with your own physician (or your obstetrician) and be guided by what he or she suggests in terms of further testing.
I hope that helps a little bit, and that I have not "blinded you with science" (which was not my intention), and I apologise for me English spelling!

TS- Dithiothreitol -DTT- Treatment of RBCs.pdfThis is our procedure for the HemoBioScience product.  it will be open for 30 days only.  (I think)
Don't worry about thawing it too many times - there is only 2-4 mls in each tube, so it doesn't last for that many pts.  We have just thawed ours at room temp.  We wrote the procedure using both the HemoBioScience procedureand the one in the AABB Tech Manual.
Best of luck

When I got to bed last night, I suddenly realised that I may have missed out a fairly obvious cause, as I had not taken into account the fact that the patients were all pregnant.

I just wonder if these patients have all made anti-Lea and anti-Leb, as it is not unusual for the Lewis antigens to "disappear" in pregnancy, and quite often they transiently make Lewis antibodies.  If none of the cells you are using are themselves Le(a-b-), then this antibody mixture (actually, it isn't a mixture, but anti-Lea+b) can look like an antibody directed against a high prevalence antigen.

If you cannot obtain sufficient Le(a-b-) red cells to test this theory (and it is only a theory), you could try to inhibit the antibody with saliva from an Le(a-b+) individual (but don't forget to control this by diluting another aliquot of the plasma with saline).
The attached may be of interest/use.
The Lewis Blood Group System and Secretor Status.docx

"Do you want it to be faster and more hands-off or more exact?"  They really said that?  Yikes!
Scott

IN the UK, the NHSBT (at least) has been performing titrations of all antibodies, of all specificities, in gel, after an extensive amount of work performed by my friend Gordon Burgess showed that there was very good correlation between these titres and those obtained by tube technique.

The age old problem of how do you make people pay attention to the details...If you figure this out, let me know. I haven't yet.
Do you not have an "IRRADIATED RBC" product in your dictionary that the physician could have chosen? That puts the responsibility on them, where it should lie. A comment is not an order and, if they are relying on that, they are forcing your techs into a position of failure. I would suggest you add an irradiated product order to your dictionary. If the physician wants that product, they must order that product that way.
 
 

Certainly the blood supplied by the NHSBT that what is on the label on the outside is GUARANTEED to be what is actually in the bag, and so no retyping is required.  I THINK the same applies in Scotland, Wales and Northern Ireland, although am happy to be corrected.
As all such "kills" would be reported to our regulatory authorities, and published in the annual Serious Hazards of Transfusion (SHOT) Report, I can say for certain that no "kills" have been reported for many, many years!

And we need to remember: patient is from Mumbai.

And we need to remember: patient is from Mumbai.

Why would the red cells of an individual who is Jk(a-b-) not react with Ulex europeaus?

Sadly, I don't (and, if I did, the seams would explode if I tried to put it on these days)!

Ah...but you have to carefully calibrate the use of that finger or fingers. And bare fingers worked better than gloved fingers. Of course, that was before we really had gloves.

We had a patient that seemingly converted from A Positive to A Negative.  We sent the patient to our reference lab and through whatever voodoo they do, discovered that she had proteins masking her D antigens.  I don't remember her specific disease process, and I don't think it was anti-D, but they reported that the patient was indeed A Positive.

The other thing could be that the clones used in the anti-D have changed, and they no longer detect a particular mutant type.

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.

Awesome responses as usual Sir!

Thanks ELondon.
Could I just say again, even if the Reference Laboratory does detect an antibody (or more than one, come to that), it is not a particularly abnormal thing in pregnancy, but it does not mean for one minute that the pregnancy will be affected; Mother Nature has seen to that.
There is another Blood Group System named Lewis.  The antigens within this system are soluble in the plasma part of your blood, and are adsorbed onto the red cells from the plasma (they are not intrinsic to the red cell membrane).  During pregnancy, the concentration of plasma lipoproteins (fatty proteins in the plasma) can increase enormously (about four-fold).  These plasma lipoproteins "mop up" the soluble Lewis antigens, and a pregnant woman, who would normally be, for example, Le(a-b+), can become Le(a-b-), and may even, temporarily, produce antibodies against the Lewis antigens (an individual hardly ever produces antibodies against an antigen that they express - but strange things happen in pregnancy!).  In addition, ALL babies are born as Le(a-b-), so any Lewis antigens Mum produces will NOT affect the baby!
There are many, many other antibody specificities that will not affect the pregnancy at all.
Now, I should say two things.  Firstly, I cannot say, from a distance, what is the antibody in your plasma (that can only be done by the laboratories at the Hospital and the Reference Laboratory, but it does not sound at all serious).  Secondly, i am what is called a Biomedical Scientist, not a doctor, and so I am, by Law, not allowed to diagnose (as far as I know, neither is the midwife), and this is why I am so glad that you are going to see an Obstetrician, who, I hope, will be able to reassure you even more.
Mean while, sleep easier, and enjoy your pregnancy!