making up an elution???

[StephanieM]

Does anyone have any suggestions on how to make up an elution unknown? I have people who need practice working up an elution, our CAP sample isn't scheduled until March and there usually isn't enough to pass around.

[Malcolm Needs ☆]

Well, the antibody doesn't have to be rare, so why not just use some D+ red cells and some anti-D?.........or isn't that what you mean?

[Dr. Pepper]

That's what I do as well for my students and trainees. That's easiest, but you could also use any outdated IgG antiserum or nice patient IgG antibody. Mix 1 cc serum and 1 cc antigen-positive packed cells, incubate 30 min at 37, check to make sure it's DAT-positive and away you go.

[AMcCord]

You can make a quick recipe of 3-4 pigtails from an O Pos donor plus 10 drops Check Cells. You might have to adjust a bit depending on the size/red cell volume of the pigtails, but this makes a nice weaker DAT sample that works well.

[PAWHITTECAR]

Thanks AMcCord that sound easy enough I think I will use it.

[L106]

Similar to how Dr. Pepper described, you can also make up a patient sample to mimic a warm autoantibody by added several drops of Anti-e to e pos red cells, incubating, then washing, etc.

For the patient's plasma, you can either use plasma that contains no unexpected antibodies, or you could add a couple drops of the Anti-e if you want the specimen to have the autoantibody in the plasma in addition to on the red blood cells.

[kirkaw]

This is so awesome that this was up for discussion today! I was just looking for a recipe for creating a positive DAT. I was going to do what Malcolm suggested. Do I have to incubate that or just combine and give to the student to test?

[Malcolm Needs ☆]

You have to incubate, otherwise the IgG antibody will not sensitise the red cells quickly enough.

[Dr. Pepper]

One thing about using reagent anti-D: most are mixes of monoclonal IgM and IgG, to react in rapid tests and weak D tests respectively. The elution procedure should work fine, but you may find the cells agglutinating spontaneously in a DAT test due to the IgM component in the reagent. Our DAT procedure is, if positive with polyspecific AHG, to repeat with anti-IgG, anti-C3 and saline as a negative control. The saline control can come out positive with cells prepared this way.

[kirkaw]

Thanks Phil. I'll remember that and do the testing before I give to the student to test.

[Malcolm Needs ☆]

I agree Phil. I was thinking about using a genuine patient's anti-D, which should be reasonably easy to come across. Of course, it doesn't need to be an anti-D. Any specificity made by a patient would do, providing that red cells expressing the corresponding antigen are available.

[Eagle Eye]

Yes. Agree. I have seen the same. But I tell my student to perform only DAT IgG once I knew this was happening. (I know cheating but they are learning elution)

One thing about using reagent anti-D: most are mixes of monoclonal IgM and IgG, to react in rapid tests and weak D tests respectively. The elution procedure should work fine, but you may find the cells agglutinating spontaneously in a DAT test due to the IgM component in the reagent. Our DAT procedure is, if positive with polyspecific AHG, to repeat with anti-IgG, anti-C3 and saline as a negative control. The saline control can come out positive with cells prepared this way.

[AMcCord]

For the patient's plasma, you can either use plasma that contains no unexpected antibodies, or you could add a couple drops of the Anti-e if you want the specimen to have the autoantibody in the plasma in addition to on the red blood cells.

This one works very well.