Positive Autocontrol in gel, negative DAT

[CMCDCHI]

We have seen a few patients recently with a positive autocontrol in MTS gel, but a negative DAT by tube testing.  The majority of my experience is in a reference lab with only tube testing and gel is new to me.  What do you make of the positive autocontrol results?  

[Malcolm Needs ★]

I wouldn't worry too much about it, to be honest CMCDCHI.

We see this quite often. It is really only the fact that gel technology is much more sensitive than tube technique, but, every now and again, gel technology is, if you like, "too sensitive". It means that the red cells are, without doubt, sensitised in vivo by either an antibody, or an antibody that has resulted in C3d coating of the red cells (or both), but it does not necessarily mean that this is clinically significant.

[yan xia]

I totally agree with Malcolm, and the autocontrol with patient plasma and cells reacted is more sensitive than DAT, I don't think it is proper to compare those two test in two differ method. I prefer do autocontrol in tube compare autocontrol in gel.

Just personal point of view.

Edited April 30, 2014 by shily

[CAMLT]

If the auto control in gel is positive, there is high chance that the other screening cells reactions in gel also positive, strength of variation may be possible. Since gel is more sensitive to warm auto, tube test including DAT may not catch. I will switch to LISS screen (not PEG) to see if screen is still positive. Sometime even DAT negative which normally not justifies to do eluate, but if you really perform the elution, there may have positive reactions across over the panel since elution tends to be more sensitive to catch warm auto. This is why to call a pt has warm, the eluate must have positive reactions as a minimum. 

 

WAA is not regarded as clinically significant, but interfering routine tests. For the future reference, you can put a comment in this pt folder to say like use LISS to do antibody screen. 

 

If WAA strong enough, auto/allo adsorption may be needed. In that case, the DAT will be surely positive. 

[Abraham]

If we repeat the autocontrol in tube and 90% of the time it is negative.

We repeat the autocontrol in tube and 90% of the time it is negative.

[David Saikin]

It would be interesting to know how strong your gel reactions are/were. I have found that a 2+ or weaker gel reaction is virtually undetectable in tube testing regardless of the enhancement. I also think it would make more sense to run your DAT in gel but that is up to you.

[Joanne P. Scannell]

I composed an answer to this yesterday and must have gotten 'sideswiped' before I posted it!

 

Anyway, here's what I have to add to this conversation ...

You cannot compare gel to tube testing.  Aside from the 'sensitivity' issue (DAT could be positive in gel, negative in tube), gel has a totally different premise that needs to be taken into consideration.

• Gel: Slippery RBCs will make it all the way to the bottom of the gel column during a given period of time at a given centrifugal force.
• Tube: Cells must be agglutinated.

1. Therefore, gel results are affected not only by coating of RBCs with antibodies, but also coating (roughening) with just about anything (proteins, medications).  In addition, changes in shape (e.g. sickling, acanthrocytosis, fragmentation) will cause a slower migration to the bottom. 

 

2. You cannot expect an auto control to give the same results as a DAT because the autocontrol has plasma in it.  (It's like all ladies are women but not all women are ladies.) Certain properties of the plasma will affect this downward migration (e.g. artifact, fibrin, TP ratio leading to rouleaux, cold agglutinins).

 

3. You cannot even expect the same 'DAT grade' with an auto vs DAT.  The autocontrol contains the same enhancement medium as the tests.  The presence of an enhancement medium will, ummm, enhance the antigen-antibody reaction.

 

Suggestion:  Don't mix your media.  If Auto Control is positive in gel, don't bother running DAT with tube method; run it using same method as the Auto Control (gel) and be sure to include a 'negative' control using a buffer card with patient's cells.

• If the buffer control is positive: You cannot determine if DAT is pos or neg because the 'positive result' may be due to steric interferrence rather than 'coated with antibody'; check hemo smear for sickling, acanthrocytes, etc.
• If the gel DAT is positive and the control is negative, your patient cells have a positive DAT.
• If the gel DAT is negative, the positive auto could be a result of plasma abnormalities, check protein analysis, cold agglutin, etc.

Hope I haven't overstayed my welcome ...

Edited May 1, 2014 by JPCroke

[yan xia]

Thanks for JPCroke 's marvellous post.

If the gel DAT is negative, the positive auto could be a result of plasma abnormalities, check protein analysis, cold agglutin, etc

I think  antigen and antibody reaction stronger  in enhanced media will also cause this kind of situation.

[CMCDCHI]

Thank you all for your input.  I won't worry too much about these seemingly conflicting results.  

 

There is a whole lot of "mixed media" in our lab.  ECHO for routine type & screen, gel for antibody ID, tube for DAT.  It makes me crazy not to have one system to rely on for one entire work-up.  Most likely, we will be phasing gel out since our contracts are with Immucor, but our techs are very attached to gel. 

[Malcolm Needs ★]

Don't worry too much about having a number of technologies CMCDCHI. As a Reference Laboratory, we rely on having multiple technologies, as this gives us the best chance of emulating the findings of the referring laboratory, but also the best chance of deciding whether or not these findings are clinically significant.

[kirkaw]

Adding onto this conversation with a slightly different spin. We had a patient with positive gel antibody screen, whose gel panel showed a perfect anti-K pattern with a positive control. The patient was transfused 3 months ago. The patient is B+. A DAT was performed and found to be negative. On a subsequent day, 2, B+, K- units were crossmatched in gel and found to be incompatible with 1+ reactions. 2 more O+, K- units were crossmatched in gel and were clean as a whistle! What gives? Should I have done an eluate?

[Malcolm Needs ★]

YES!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!

[subbusp58]

I need a format to record and classify  near miss events that occur in sample collection, intralab testing  and events during blood administration. if someone could provide one such document, I would be thankful.

[mollyredone]

Thought  I would resurrect this topic.  I've been trying to find a definitive answer to my question regarding positive autocontrols and what type of DAT should be done.  We do our panels and AC in gel.  I decided that the DAT should be done in gel, using IgG.  Is this the right approach?  If we have a positive DAT, we usually send it to the reference lab for workup.

Thanks!

[Malcolm Needs ★]

I would certainly do it in gel, but I would use the cards that contain monospecific reagents for anti-IgG, anti-IgM, anti-IgA, anti-C3c and anti-C3d.

Edited February 26, 2016 by Malcolm Needs
Spelling - again!

[mollyredone]

We only stock anti-IgG cards.

[amym1586]

We only have the IgG cards and we don't do eluates.   Makes you feel kinda stuck.

[SMILLER]

We do antibody IDs and screens in gel, but switch to tube when we need to do a poly DAT.  We do not maintain compliment QC however, so if we do get a positive DAT that has to be resolved, we have to send it to another Lab.

Scott

[mollyredone]

So, if we have a positive autocontrol, the DAT should be performed by tube with Poly reagent?  If it's positive, we would use separate IgG and complement reagents.

ETA:  This is what we do for DATs for hemolytic anemia

Edited February 26, 2016 by mollyredone
didn't want to sound stupid

[SMILLER]

That's what we do.  With a positive poly (anti-IgG and anti-compliment), you have to differentiate the reaction. 

Scott

[amym1586]

If the DAT is positive, we perform poly in tubes.  If the poly is Positive we'll perform the IgG.

[StevenB]

First and foremost...if you are going to go back and do a DAT because the AC came up positive, make sure you do it with a freshly made cell suspension.  Also, if taking that step and you have the reagents, test with polyspecific (PS) and IgG at the same time  since you suspect it will be positive.  If you only test with PS and it comes up positive, you'd have to make ANOTHER fresh suspension to test with IgG if you were going to do that.  The 10-12 minutes it takes to do the PS test is too long for cells to sit in suspension for an accurate IgG DAT.

[Dansket]

I never stocked polyspecific reagent, just anti-IgG and anti-complement reagents separately.  Test adult cells with both if physician requested DAT on adult and when working-up patients with autoantibody.

[Dansket]

4 hours ago, StevenB said:

First and foremost...if you are going to go back and do a DAT because the AC came up positive, make sure you do it with a freshly made cell suspension.  Also, if taking that step and you have the reagents, test with polyspecific (PS) and IgG at the same time  since you suspect it will be positive.  If you only test with PS and it comes up positive, you'd have to make ANOTHER fresh suspension to test with IgG if you were going to do that.  The 10-12 minutes it takes to do the PS test is too long for cells to sit in suspension for an accurate IgG DAT.

Another advantage of automated testing, all cell suspensions are freshly made.

[Malcolm Needs ★]

10 hours ago, Dansket said:

Another advantage of automated testing, all cell suspensions are freshly made.

Not quite.  Your A₁ and B reverse grouping cells are not.