Antibody stimulation by antigen negative blood?

[RichU]

I PUT THIS Q IN TRANSFUSION SECTION BUT I GUESS MORE RELEVANT HERE. 

We recently had an antenatal case which showed some unusual features.

This was the third pregnancy, the first miscarried at less than 10 weeks. The second was delivered after a seemingly uneventful pregnancy.

At booking we found enzyme anti-f. At 28 weeks anti-f, anti-K and a further unidentified Ab. (f and K enzyme only).

By 32 weeks She also had anti-Fya and possibly Cw.

From 38 weeks gestation we kept 4 R1R1 K- Fya- units crossmatched in case of emergency. These were compatible.

4 days later the sample had a positive DAT and all units were incompatible. The patient delivered before we could get the sample referred to the NHSBT lab in the UK.

The baby typed as R1r K-. The baby had a weak positive DAT so Fya antigen typing was not officially performed ( I did the test and am certain It was Fya-)

So, my questions are; How can the anti-K and anti-Fya be stimulated by this fetus? Has anyone else seen this happen?

I was hoping to present a simple(ish) talk on anti-f to the multi-disciplinary on-call team at my hospital but don't want to scare them!

Thanks,

Rich

[Malcolm Needs ★]

How did you type the baby for the Fy(a) antigen, if the DAT was positive?

[RichU]

Unofficially!

IAT gelcard. Result 1+ was weaker than DAT 2+ and control Fya+b+ cell 4+.

Even though Fya type not valid the K antigen type is.

[Malcolm Needs ★]

Thanks.  I'll have a think.

[John C. Staley]

30 minutes ago, Malcolm Needs said:

Thanks.  I'll have a think.

Malcolm, I'm glad to see you are on the case.  When I first read the question in the Transfusion section I knew I wasn't going to touch it!!  Looking forward to see what you come up with.    

[Neil Blumberg]

Probably the primary stimulation was not the fetus, but previous pregnancy, transfusion, tattooing, needle sharing, non-sterile tattooing, etc.  Pregnancy is a situation in which B cells are upregulated (type 2 immunity) and T cells down regulated (roughly speaking) and pregnancy may have increased B cell activity to the point where previously undetectable antibodies are now detectable.  Just a theory ;).

[RichU]

Thanks.

I did hypothesise that stimulation of her immune system by current pregnancy triggered production of antibodies made in response to previous baby which were sub detectable at booking.

ie. Patient made anti-f, anti-K and anti-Fya when previously pregnant, current fetus (f+, K-, Fya-) lead to stimulation and increase in production of all 3 antibodies by general triggering of immune system.

Not sure if this occurs and didn't want to float that idea straight away.

Rich

[Malcolm Needs ★]

I have been thinking about this and I have come, more or less, to the same way of thinking as Neil Blumberg.

The first pregnancy almost certainly could not have caused sensitisation of most of the common antigens, as some would not be formed on the foetal red cells at 10 weeks of gestation, while, even with a huge foeto-maternal haemorrhage (FMH) (in terms of the ratio of the total foetal red cell mass), the actual volume of foetal red cells transferred to the maternal circulation would not be sufficient to cause sensitisation in anyone but a person who is a "super responder".

The second pregnancy could easily have caused a primary response, either at partum, or as a result of a chronic FMH throughout much of the pregnancy.  Unless the woman's antibody screen was performed at about six months post-partum, such antibodies may never have been detected at their peak, and then may have declined to levels where normal serological techniques would not detect them.  Of course, all of this is theoretical, but, given the fact that the mother is probably an R1R1 (from information given above), it is most unlikely that an FMH estimation, such as a Kleihauer test was performed, or that the mother would have been serologically "followed up".  You also do not give the woman's transfusion history.

It would be helpful to know the time between the second and third pregnancy, and also, of course, if the same male was the biological father in both pregnancies.  This latter piece of information may actually be very difficult indeed to ascertain, as the woman may not wish to disclose her sexual history.

It is not unknown for antibodies of a particular specificity (say, purely for example, an anti-Jka) to increase in strength (as measured by serial titres) during a pregnancy that is, in this example, Jk(a-).  This is more common in a pregnancy where there is at least one other specificity (let us say, again, just as an example, anti-K), where (again, just as an example) the foetal red cells do express the K antigen, but not the Jk(a) antigen.

Lastly, antibodies that are forming de novo, very often seem to cross-react with antigens to which they are not actually stimulated.  This is true, even in the case of some monoclonal antibodies (particularly those within the HLA system), but some antibodies (again, even monoclonal antibodies) maintain what I will call a pseudo-specificity even in the "mature state".  This includes monoclonal anti-D.  Thorpe et al have reported that monoclonal anti-D molecules possess a V4-34 moiety, that is also present in anti-I and  anti-i, which is why these reagents should never be used straight from the fridge (Thorpe SJ, Boult CE, Stevenson FK, Scott ML, Sutherland J, Spellerberg MB, Natvig JB, Thompson KM.  Cold agglutinin activity is common among human monoclonal IgM Rh system antibodies using the V4-34 heavy chain variable gene segment.  Transfusion 1997; 37: 1111-1116 and Thorpe SJ, Ball C, Fox B, Thompson KM, Thorpe R, Bristow A.  Anti-D and anti-i activities are inseparable in V4-34-encoded monoclonal  anti-D: the same framework 1 residues are required for both activities.  Transfusion 2008; 48: 930-940).

I hope this helps a little.

[RichU]

Again thank you.

The phenotype is probably R1R2 which is why I opted for anti-f and not anti-c.

No transfusion history.

The second pregnancy delivered in March 2018. We had no sample after her 28 week routine A/N in Dec '17.

The booking sample in which we found anti-f was in June '19 (approx 10 week gestation)

The woman's partner in the latest pregnancy is K- but we know how much store to set by that!

49 minutes ago, Malcolm Needs said:

It is not unknown for antibodies of a particular specificity (say, purely for example, an anti-Jka) to increase in strength (as measured by serial titres) during a pregnancy that is, in this example, Jk(a-).  This is more common in a pregnancy where there is at least one other specificity (let us say, again, just as an example, anti-K), where (again, just as an example) the foetal red cells do express the K antigen, but not the Jk(a) antigen.

I will go with this.

Rich

[Arno]

Is she given plasma derived product (not talking here about anti-D prophylaxis)? Thinking more here about IVIG?

[Malcolm Needs ★]

4 hours ago, Arno said:

Is she given plasma derived product (not talking here about anti-D prophylaxis)? Thinking more here about IVIG?

Good point.

[RichU]

Hi Arno,

I am unsure about this.

Why might she have been given IVIG and what effect could this have on the serology?

Cheers,

Rich

[Neil Blumberg]

IVIgG can contain detectable rbc alloantibodies, although it would be unusual to be anything other than anti-A and anti-B.

[Malcolm Needs ★]

True, but, in the early days, I know of at least one example that caused almost pure red cell aplasia in a BMT from a group A donor, because the contaminating anti-A was so strong.

[Arno]

Hi Rich,

I am not a clinician but as far as I know IVIG can be given to obstetrical patient in diff. conditions (autoimmune disorders, recurrent pregnancy loss, ...).

I thought about IVIG when I saw the DAT becoming positive plus additional reactions coming up over the time. Anti-A and Anti-B are indeed the most prevalent antibodies in plasma derived products but other specificities of low titre can be present sometimes such as anti-D, anti-K and a bunch of antibodies of undetermined specificity reacting with several to not say all RBCs. 

Just a thought that can be doublechecked with the clinician..?

Hereunder is a very great (not recent though) paper to be read and re-read again:

Problems Associated With Passively Transfused Blood Group Alloantibodies 

George Garratty, PhD, FRCPath

American Journal of Clinical Pathology, Volume 109, Issue 6, 1 June 1998, Pages 769–777, https://doi.org/10.1093/ajcp/109.6.769

[mrmic]

I certainly agree with Mr. Blumberg and Mr. Needs as well as others, everyone brings up excellent points and explanations.   My only comment I could put forth for consideration would be from a BB Pathologist I once worked with many years ago having observed similar cases.  "Pregnancy is a disease".  

[Malcolm Needs ★]

I've had another thought about the baby's Fy(a) type being 1+, even though the control was 4+.  It could be that the maternal anti-Fya was strong enough to be a blocking antibody.  I have grave doubts about this, as blocking is rare, and I have never heard of a blocking anti-Fya, but there is a first time for everything!

[RichU]

Thanks for still cogitating on this.

How strong would an antibody have to be to block antigen sites? ( Difficult to answer given the lack of cases, I assume)

Also, given the presumed whoppingness(!) of the titre, wouldn't you expect at least some sign of HDFN?

Cheers

p.s. I have had to postpone the presentation due to circumstances beyond even World leaders.

Edited May 15, 2020 by RichU
addendum

[Malcolm Needs ★]

Actually, you are quite right; there would have to be signs of HDFN - so forget the idea of a blocking antibody.  I was letting my imagination run away with me.

SORRY - silly idea!

[mrmic]

Anti-Fya and Anti-Fyb are not well known to cause significant HDFN.   I have not seen one, at least.  Was there any follow-up testing of the infant?  What were the laboratory findings, i.e. bilirubin etc. ?  It has been a few months now, have you had a chance to re-type the infant's red cells?   Is there a chance that it really was a weak binding of Anti-Fya with Fya+ red cell antigens?   If only a gel-card method of interpretation of a "weak positive" as being negative was used, I wouldn't necessarily be convinced that the infant is Fya-.   Gel card methods do funny things sometimes.

[Malcolm Needs ★]

28 minutes ago, mrmic said:

Anti-Fya and Anti-Fyb are not well known to cause significant HDFN.   I have not seen one, at least.  Was there any follow-up testing of the infant?  What were the laboratory findings, i.e. bilirubin etc. ?  It has been a few months now, have you had a chance to re-type the infant's red cells?   Is there a chance that it really was a weak binding of Anti-Fya with Fya+ red cell antigens?   If only a gel-card method of interpretation of a "weak positive" as being negative was used, I wouldn't necessarily be convinced that the infant is Fya-.   Gel card methods do funny things sometimes.

While I would agree with you that no antibodies within the Duffy Blood Group System are well known for causing clinically significant HDFN, and repeating what I said above, that I have grave doubts as to this being such a case, the fact that you have not seen one does not mean that such things do not occur.  I would cite Goodrick MJ, Hadley AG, Poole G.  Haemolytic disease of the fetus and newborn due to anti-Fya and the potential clinical value of Duffy genotyping in pregnancies at risk.  Transfusion Medicine 1997; 7: 301-304 (doi: 10.1046/j.1365-3148.1997.d01-38x), and indeed there has been a report of a "blocking" anti-Fya (which I should have remembered, not least because four of the six authors are personal friends!), namely Lee E, Cantwell C, Muyibi KO, Modasia R, Rowley M, New H.  Blocking phenomenon occurs with murine monoclonal antibodies (anti-Fya) in a neonate with a positive direct antiglobulin test due to maternal anti-Fya.  Blood Transfusion 2015; 13: 672-674 (doi: 10.2450/2015.0232-14).

I would query your choice of words with regard to, "Gel card methods do funny things sometimes."  I would rather like to know exactly what you mean.

[RichU]

Sorry for the delayed reply mrmic.

We received 3 requests for bilirubin level but 2 were insufficient. The first was within normal limits.

We have had no follow up samples from baby and I do not expect one.

I assume, due to our lack of knowledge regarding an ongoing situation, that the baby did not require any medical intervention.

Thanks

[mrmic]

Wow this is a late post.   I just can't find the time to keep up sometimes.

I certainly was not implying that either Duffy antibody would not be able to cause HDN but rather theoretically speaking given the circumstances it didn't quite give the picture of HDN.   Again, even that is not a absolute.   Looking back at all the comments and possible causes, which all had merit,  I failed to see any reference to the possibility of an autoimmune issue and that there may be a possibility that the specificities are part of an newly development of autoantibody complex forming, i.e. mimicking specificities.   Although these are normally seen within the Rh-Hr specificities, other specificities are not unheard of.   Follow-up testing for cases like this rarely pan-out, if the infant clinically unaffected, the parents get their baby and disappear (sometimes and at least may not show up again until the next pregnancy).  Too bad, would make a good abstract....

"My" thoughts or opinions for this site are based on previous experiences or readings (actual book in hand journals) and etc.    Immunohematology Reference Laboratories see  a variety of cases sent for consultations and that is what makes it so intriguing and challenging for us to give the clinician the information he/she needs to take care of their patient and that we are right there with him to help.  We may not always have a specific answer but we can look for histories of similar cases and what the outcomes have been and give it our best educated interpretation of what might be happening and what transfusion recommendations we might propose.

I'm about to retire and my ramblings will decrease (Yea goes the crowd). 

As far a the gel system, again my own thoughts/experiences we had in our Immunohematology Reference Lab, starting back even  before Ortho commercially prepared system was as follows:   Basically it is a micro-LISS-system with an optimized serum to cell ratio.  Although we could not find a niche for using it on our investigations, we did start keeping it around to reproduce issues our hospitals were seeing with its use their routine transfusion service and to help provide educational information on what was happening and whether it had any clinical relevance.   There was a lot of weak reactivity of various strengths referred to us by a variety of hospitals.  Many these were related to the problems seen with  the LISS tube system.  Maybe even a little more since it much more sensitive based on how the method is set up commercially to work.   

Lastly, I believe that Malcolm Needs is truly an asset to this site and provides excellent information to all regarding such a variety of topics and also provides excellent references to support the information he provides.   Thank you Mr. Needs!   I hope you continue to provide your insight in this forum for many more years.

mic

 

[RichU]

I used this case study as part of my Higher Specialist Diploma in Blood Transfusion.

The IBMS have asked if I would like to give my PowerPoint presentation ('What the f?') at the 2023 Congress.

Thank you to all the contributors - I will certainly big up PathLabTalk if I do get to do it.

Rich