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Unusual Finding

PTT Mixing Study Prolongs Instead of Shortening

This is a prior-year post that has resurfaced as we have switched over to our new provider and format. Here is another interesting question about mixing studies from blog participant Sue Hollister:

Over the past year we have had a few aPTT mixing studies where the initial 1:1 mix was actually longer than the original prolonged aPTT.  For example, the original prolonged aPTT was 40 seconds, the initial 1:1 mix was 43 seconds and after 1-hour incubation, the mix was 45 seconds. We are using a commercial normal pooled plasma to perform our mixes and this has happened with both fresh and frozen samples.  We have ruled out heparin and other anticoagulants as well.  Do you think it has something to do with the draw itself?
Thank you, Sue Hollister

Hello, Sue. This is the kind of occurrence that is never discussed in the procedure or the package insert, so it leads to some “scientific”  speculation. The first assumption is that no heparin or Coumadin is present to prolong the PTT, and you’ve confirmed this.

Next, determine whether the prolongation is within the CV% (precision) of your PTT. If so, it has no significance.

The key artifact that affects mixing studies is the presence of platelets or platelet products in the patient and the normal pooled plasma control. Make sure your patient’s plasma platelet count is centrifuged well and consistently <10,000/uL. Though platelet secretions typically tend to shorten the PTT, they could be releasing a weak inhibitor.

Likewise, ensure your reagent normal pooled plasma is platelet-poor. If you are purchasing it from a supplier, as most do, check with the manufacturer to confirm. Most prefer frozen to lyophilized plasma to avoid a matrix effect.

Finally, if the inhibitor is real, it may be weak and its effect may be developing during the short time the plasma stands while preparing the mix. While the books say lupus anticoagulant is revealed in the immediate mix and anti-factor VIII develops upon incubation, there is a crossover. Further, you could be dealing with a weak inhibitor to another factor, such as anti-V, which arises subsequent to the use of fibrin glue.

If a weak and slow-developing inhibitor is the culprit, you may want to try a 4:1 mix of patient plasma to normal pooled plasma control.

I hope this helps. I’d be happy to hear how others deal with this problem. Geo

Comments (2)
Nov 1, 2022 9:09pm

We’ve seen this pattern a few times with a strong LA; we even confirmed this pattern in an external quality assessment exercise using the RVVT (Favaloro EJ, Bonar R, Zebeljan D, Kershaw G, Marsden K. Laboratory investigation of Lupus Anticoagulants: mixing studies are sometimes required. J Thromb Haemost. 2010 Dec;8(12):2828-31. doi: 10.1111/j.1538-7836.2010.04090.x. PMID: 21029364.

Mar 2, 2009 6:17am

An initial prozone effect of patients ptt with dilution of t
An initial prozone effect of patients ptt with dilution of the phospholipid antibody in the 1:1 mix should be considered.

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