Gel card with 1+ reactions showing negative after spinning a second time

[Cornelia Kuehn]

An ER trauma patient's plasma showed 1+ reactivity in Gel with Screening cell 1 of Immucor's Trio screen. The other two cells were negative. (We buy 3% Immucor screening cells, then dilute them to 0.8% for gel testing.) Using an Ortho 0.8% panel for antibody identification, we found 7 of the 11 cells reacting 1+ with a negative auto control. The reactions did not fit any pattern what-so-ever. Our workup was stopped at that point because the patient was transferred to a larger facility. On follow-up the next day, we found that the other facility had gotten a negative antibody screen, also using Gel cards. Within three hours of performing the antibody screen on the ER trauma patient, the remaining three wells of the gel card were used to perform an antibody screen on another patient. So, the card with the positive Screening cell 1 was spun for another 10 minutes. After the 2nd spin, the reaction had changed to negative. Later, we spun the two cards containing the antibody id with the seven 1+ positive reactions. After the 2nd spin (for a total of 20 minutes) most of the reactions were now negative with the others only questionable, definately not 1+ as after the orginal spin.

I've checked the rpms and timer on the centrifuge, and they are okay. Has anyone else seen positive reactions change to negative after spinning a second time?

the problem is if you spin a second time you change the method by itself and you get a result that you cannot validate..... but I think you have to look if the patient have autoantibodys it seems like you have to eluate and see what is on the surface of the ery`s

[geekay]

Hi all,

I fully agree with Dr Susannah....

This definitely suggests that "rouleaux formation" does give "false positivity " in the gel technique... I had many similar experiences with Diamed few years back...

Though I had brought up the topic some time back in this column, it went unnoticed anyway....

This querry, when it was raised to the "Diamed company" few years back, nobody from the company could give any explanations in this regard.

This is a warning signal for all who blindly follow the "gel techniques" !

best wishes....

[Liz]

Spinning a Gel card reaction twice invalidates the original reaction.

I totally agree. That is why there are instructions.

LIiz

[drsbright]

NOWHERE does it state that it is acceptable to respin the gel cards. How did this get started? That cannot possibly be acceptable practice and these results should be tossed out.

drsbright, Indianapolis, Indiana

[LaraT23]

Personally I would have thrown them in a tube and saline replaced if necessary. Especially in trauma patients, who knows what type of medications have been given and also what type of actue phase reactants were in the plasma. True agglutination will show in the tubes and will still be there even if you saline replace. I would scope the tubes, as the gel is more sensitive.

[NedB]

The ProVue examines every card before accepting it for testing; sometimes the buffer just above the gel forms bubbles, probably from rough handling. We spin these cards before use and the ProVue invariably accepts these.

As for the re-spinning - not a good idea. Re-spinning the plasma from the sample (preferably in a micro-centrifuge - 15,000+rpm) and investigating for rouleaux are better courses. If the rouleaux does not show, double the volume of plasma and re-check; remember the gel is ultra-sensitive.

Question for Susannah: Do you have the product insert for the Bio Gel? I evaluated it way back but didn't keep the literature. Who knew?

[n.peters]

A couple of years ago an evening shift tech where I worked got a 1+ weak reaction and decided to respin the card. The respin was negative and the tech reported it out as negative. Some how day shift caught this and repeated the test. It came out as anti-E.

[NedB]

I did not have direct evidence like you did; but it what I would have expected. I rest my case.

[KECIA]

An ER trauma patient's plasma showed 1+ reactivity in Gel with Screening cell 1 of Immucor's Trio screen. The other two cells were negative. (We buy 3% Immucor screening cells, then dilute them to 0.8% for gel testing.) Using an Ortho 0.8% panel for antibody identification, we found 7 of the 11 cells reacting 1+ with a negative auto control. The reactions did not fit any pattern what-so-ever. Our workup was stopped at that point because the patient was transferred to a larger facility. On follow-up the next day, we found that the other facility had gotten a negative antibody screen, also using Gel cards. Within three hours of performing the antibody screen on the ER trauma patient, the remaining three wells of the gel card were used to perform an antibody screen on another patient. So, the card with the positive Screening cell 1 was spun for another 10 minutes. After the 2nd spin, the reaction had changed to negative. Later, we spun the two cards containing the antibody id with the seven 1+ positive reactions. After the 2nd spin (for a total of 20 minutes) most of the reactions were now negative with the others only questionable, definately not 1+ as after the orginal spin.

I've checked the rpms and timer on the centrifuge, and they are okay. Has anyone else seen positive reactions change to negative after spinning a second time?

Hi, we see this alot and have found on several occasions that if we seperate(from the red cells) and re-spin our plasma or even repeating it in tube with albumin that these unexplained reactions pan out to absoluting nothing but a wasted of time.

[Eoin]

Wow, now here is a thread that has stayed the distance!!! I have been using Gel cards since first released many many years ago. That was in a hospital in Oz with large trauma and maternity centres. I have looked at this question a number of times (at others requests)> NOWWHERE in the literature does it say re-spin.

Initial inspection of cards on receipt will show bubbles, or cards that have been shipped on their side. It is permissable to spin these to re-settle the gel. BUT it will not get rid of all nuisance reactions. Colloids and other plasma expanders would be one of my chief suspects (along with other s&%$) they pump in for ER or obstetric emergencies. The others like fibrin etc can be dealt with easily, but does mean a repeat. The others - Good Luck with all that. Experience is essential!! Remember, you can treat a patient with a reaction, but cannot help one who is exanguinated! Eoin

[ABADI]

We have also seen positive reactions spinning down to negative when respinning the plasma again, the gel card while finishing a second screen on a new patient on the same card. We have found that anything truly positive does not spin away. The positive reactions we had that spun away were either rouleaux or a non specific cold agglutinin. We have also found that any agglutination, no matter how little, is an antibody. Even some times with Antibody screening , the same issue

[donellda]

We have seen this problem quite a bit and began to suspect that it was a problem with the prediluted screen cells being out at room temperature for too long. We started rotating 2 sets of cells. At the beginning of each shift we take a set out from the fridge and return the set that we were using back to the fridge. This way they are not at room temperature for longer than 8 hours at a time. It seems to have helped. Last week I received a letter from Ortho discussing this same problem. They recommend that you do not store your cells near light and near an area where heat is generated. We still get the odd weak reaction that ends up being a non specific reaction but we get it a lot less.

[jlemmons]

We have had this problem quite a bit. We have two ProVues and will repeat the patient on our second machine and usually get a negative. If the second one is positive we will do a panel. Sometimes we will get all panel cells negative so we Coombs Crossmatch the units and call it a TWID (too weak to identify). These usually do not repeat themselves in the future specimens.

[irshadaad]

it hapens several times with us that the ABORH-newborn card gives all tues positve.....and on pretest centrifugation we get a normal result...we found that during transportation if the cards get disturbed due to fall or tilt..the solution layer above the gel is no more appearant in that case the card shows a fase result after centrifugation the liquid layer is restored and the cards dont show any abnormality....so its now practice with us to spin the cards before use ....we dont know if it has some logic or not

[noelrbrown]

You have to be careful diluting 3-4% cells down to 0.8% becasue of the diluent. Generally manufacturers supply their 3-4% cells in alsevers solution or a diluent that is normal ionic strength. 0.8 % cells are in a special diluent formulated for a gel card which is low ionic strength. If you do dilute cells for use in a gel card make sure the manufacturers diluent is used to resusepnd the 0.8% cells

[ewaibel]

did the cards look like they were at the correct liquid level to begin with? did you see any splashing in the cards prior to first spin? was the liquid on the bottom of the gel instead of the top? all of these could lead to an incorrect result on first spin.

[Malcolm Needs ☆]

You have to be careful diluting 3-4% cells down to 0.8% becasue of the diluent. Generally manufacturers supply their 3-4% cells in alsevers solution or a diluent that is normal ionic strength. 0.8 % cells are in a special diluent formulated for a gel card which is low ionic strength. If you do dilute cells for use in a gel card make sure the manufacturers diluent is used to resusepnd the 0.8% cells

noelrbrown makes a very important point, which many people easily forget.

[irshadaad]

no we dont use any other diluent other than supplied by diamed diluent 2 thats LISS ,the problem comes due to transportation disturbance not due to diluent....thanx for the information

[eric1980]

This topic was created a couple of years ago... -_-"

But anyway, all research and tests are done by the manufacturer on the first spin only. So when something comes up on the first spin, it means that there is something, and respinning in the hope of it becoming negative is... somewhat a denial feeling to me. =Þ

I was actually surprised that respinning of the gel cards is actually a possibility.

But I've learned from here that if the gel card is weakly positive, I should check for rouleaux formation.

[galvania]

The point about not spinning the cards a second time is relevant ONLY after the cells have been added. Irshadaad is absolutely correct that sometimes during transportation, if the cards are mishandled, the liquid can 'jump' to the cuvettes at the top of the card (the cards would have to be upside down for the liquid to be at the bottom of the well) and this can give false positives due to contamination. In this case, it is absolutely correct to spin the cards prior to use to bring the liquid back down on to the gel. However, Irshadaad, if tis happens regularly, you should contact your distributor ad try to find out where the problem lies

[Eoin]

it hapens several times with us that the ABORH-newborn card gives all tues positve.....and on pretest centrifugation we get a normal result...we found that during transportation if the cards get disturbed due to fall or tilt..the solution layer above the gel is no more appearant in that case the card shows a fase result after centrifugation the liquid layer is restored and the cards dont show any abnormality....so its now practice with us to spin the cards before use ....we dont know if it has some logic or not

We also had the same problem some years ago. We now routinely spin on receipt (could be small trapped bubbles due to disturbance in transport) and store upright - Problem solved.

Also where is the authority to spin a second time when reading coming from??? Never seen that in the literature. -

[Malcolm Needs ☆]

We also had the same problem some years ago. We now routinely spin on receipt (could be small trapped bubbles due to disturbance in transport) and store upright - Problem solved.

Also where is the authority to spin a second time when reading coming from??? Never seen that in the literature. -

Neither have I, and this is something that has worried me from the start of this thread.

[galvania]

Also where is the authority to spin a second time when reading coming from??? Never seen that in the literature. -

Cards must NEVER be spun a second time when reading. See previous comments on this thread

[Brenda K Hutson]

Yes...we've seen this phenomenon at our facility as well. Am not certain if it's a problem with centrifugation or if it could be remedied by pre-spinning the gel cards prior to use. I vaguely recall some facilities having pre-spun the cards on receipt but never got an explanation of this practice. Upon getting a positive reaction on the 3-cell screen, I always set up a new card and repeat prior to running a panel to avoid this problem. Have not seen anything from Ortho to suggest that others have this problem and/or explanations to this.

Also...I like the Immucor cells myself, and do often reduce them to 0.8% and run them in gel as you do ... does Immucor sell 0.8% panels? If not, they should...have found their cells to have a stronger antigen expression than the Ortho cells.

I mentioned to an Immucor Rep. once, that they could expand their business with "Gel users" by making 0.8% Panels also. He indicated that this would appear to be a conflict of interest; that it would be kind of like promoting GEL over their methods.

I personally don't think it is going to influence the method people use, one way or the other. And it sure would be helpful to not have to dilute the 3%!

Brenda Hutson, CLS(ASCP)SBB

[kelliott]

You should not spin a second time. You will get false negative results doing that and in addition that action is not following manufacturers directions. You should go back and look for the antibody causing the 1+ reaction on your screen cells, rather then trying to get rid of reactions. We found that Ortho's prediluted cells have a high degree of specificity but also a high degree of sensitivity. If we suspect a problem with the ortho cells we work with a 3% panel diluted to 0.8%.