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Just wondering whether anyone else has been having problems "recently" with Eluate Testing performed in GEL?  We have had a number of instances now where all cells are reactive; but if we repeat the Testing by Tube, we get specific reactions.  I have already tried having staff do the "final" spin of the eluate in a  centrifuge with a harder and longer spin....but still having problems.  Not sure if there are any current problems with Immucor Eluate Kits and/or Ortho GEL Cards and/or "just us?"

Would simply switch to Tube except that currently, we only receive the 0.8% Panels (and don't want to waste those cells) so would have to add a 3% Panel.

 

Thanks,

Brenda Hutson, CLS(ASCP)SBB

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Our last elu-kit went wonky on us and gave us eluates that reacted with everything we tested in gel. It did likewise by the tube method except I think one selected cell was negative. We discovered it on our last survey. When the second survey sample did the same thing we tried a new kit and got a specificity so we knew it was the kit.  Too late for the first sample and couldn't get more.  The kit had been open a long time so we figured it got contaminated but it didn't look bad. Made sure no patients had been affected since it last worked right so we were okay.

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We have seen similar problems at our facility Brenda.  Our last CAP survey did just as you described.  after reading the Survey Discussion, this problem was widespread and caused CAP not to score that sample.  Investigation revealed manufacturing issues that are to be corrected before the next survey.

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We had a recent rash of invalid results because the last wash control came up positive (have never, ever seen this in the past...and the eluates on the patients were either negative or showed a clear & expected specificity).  We threw out the wash solution two times within a few days because we suspected possible contamination;  finally switched to a brand new bottle to store it in as well as just happened to switch to a new lot number of Ortho screening cells about the time of the second switch.   Anybody else have this issue?

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Ah....now we get to the CAP Survey!  Are you all talking about the one that was supposed to be Anti-e?  Because for us, all cells reacted.  But our problem was not just with that survey; we had some patients react like that around the same time (but not with Tube Eluate Testing). 

I had already tried:  Making new Wash Solution; Centrifuging the final Eluate in a different centrifuge (harder, longer spin).  Was less suspicious of Eluate Kit than GEL Cards since Tube Eluate Testing results were acceptable.

Thanks for all of your input!

Brenda

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We had same problem with last CAP survey. We actually ran the eluate with ortho's ficin (gel) and the 3 cell came up negative showing anti-e specificity.  Then we ran the eluate with orthos A panel (gel) and showed anti-e specificity. I just thought that our C panel's 3 cell some how got contaminated, but after talking to friends at neighboring hospitals I found out that we were not the only ones that saw this problem.

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Interesting that the survey they didn't grade was not the sample that had anti-e.  

 

We had false positives with the modified tube method as well as with gel with the # one eluate. When the Kidd antibody in specimen #2 reacted with everything too, we used a new Elu-kit and got a clear Jka (I think) that reacted fine in gel--same cards as far as I know.  We had a student test a sample a few weeks earlier with the Elu-Kit and get everything positive but we just thought it had a bit of an autoantibody (we did not really need to test it but thought it would be interesting for the student).  The eluate on a patient before that (a couple of months) using the same kit showed a normal specificity so we concluded that the kit was defective.

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Just wanted to bring this issue up again (as we continue to experience problems).  Here is the dilemna:

1.  Reactions of Eluates in GEL that have a hazy, grainy look throughout GEL, sometimes mixed-field appearance also.

2.  Often also "appears" to be hemolysis in top of GEL well (above the GEL itself)

3. Does not occur in every patient

4.  Most of the time, these are repeat Negative by Tube Eluate Testing

 

What I have tried so far:

1.  Having them make up new Wash Solution

2.  Doing the last spin (where it is just the eluate in the tube and you are getting rid of any exraneous cells) of the eluate in a centrifuge with a higher speed and longer time

3.  We have already been checking the pH of the eluate; not just relying on the blue color

 

Any other thoughts/ideas/suggestions? :wacko: 

Thanks,

Brenda

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I don't have any experience with this kit but I would guess that there may be an incompatibility between the buffers used to preserve the CAP samples and/or the elukit and/or the reagents you are using

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Brenda:

 

I had the same problem ~2weeks ago. I'm currently using Gamma Elu-Kit (Lot:359044). Patient with a history of autoanti-E showed warm on gel panel and AC was positive. We then performed gel panel from eluate and reaction was strong 4+ on all cells(no mixed field). Repeated tube w/PEG and sure enough, anti-E came through.

 

As a result of this we had to discontinue routine use of gel DAT's, Elutions.

 

Hope this provides some consolation that you are not the only one..

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Actually mprado, as your free antibody was an auto-anti-E (which, in all probability was an auto-antibody that was "mimicking" an anti-E, as far as specificity is concerned, i.e. it could probably have been adsorbed to extinction with E- red cells), your results by EluKit of a panreacting antibody in the eluate were very probably genuine, as the elution would have contained a "concentrated" version of the auto-antibody. This "concentrated" version of the auto-antibody would probably have shown less specificity for the E antigen, and more general specificity (or non-specificity, if you like), and there was probably nothing wrong with your kit, or performing the elution tests in gels.

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On 6/24/2013 at 1:00 PM, Brenda Hutson said:

Just wanted to bring this issue up again (as we continue to experience problems).  Here is the dilemna:

1.  Reactions of Eluates in GEL that have a hazy, grainy look throughout GEL, sometimes mixed-field appearance also.

2.  Often also "appears" to be hemolysis in top of GEL well (above the GEL itself)

3. Does not occur in every patient

4.  Most of the time, these are repeat Negative by Tube Eluate Testing

 

What I have tried so far:

1.  Having them make up new Wash Solution

2.  Doing the last spin (where it is just the eluate in the tube and you are getting rid of any exraneous cells) of the eluate in a centrifuge with a higher speed and longer time

3.  We have already been checking the pH of the eluate; not just relying on the blue color

 

Any other thoughts/ideas/suggestions? :wacko: 

Thanks,

Brenda

Reviving this. We are trying to validate the Hemobioscience EluClear in gel method and we are seeing 1, 2, 4, however our last wash is clean (no hazy/grainy look, no appearance of hemolysis on the top of the gel). Any suggestions? 

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You have to spin the final eluate very hard and for some time, try 5000 RPM for 5 minutes in a benchtop.  We have validated our kit in Gel card systems.  Other manufacturers kits may or may not be validated in a gel card.  I suggest you read the IFU carefully and or call the manufacturer technical support. 

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Ensure your acid eluate kit is approved for use in the specific gel you are using- Immucor eli kit is not approved for use in gel. Ortho has a kit specifically approved for use in their gel( not others)- gel is not all the same- different matrix’s/ composition. 

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