Mixing Studies: 4:1?

Here are two new questions from “mirnaibarra” in El Paso.

George, Thank you for your prompt answer to my last question. We are setting up mixing studies for PT and PTT and I found some literature that states that a 4:1 mix is more sensitive than the 1:1 mix. Do you recommend the 4:1 mix to be used routinely as the initial or only mix, or is the 4:1 recommended only if equivocal results are obtained with the 1:1 mix?

Also, what is the most common and preferred way of interpreting the corrections. A percent correction, a percent of PNP or the Rosner Index?

Hello, and thank you for your questions. Most coagulation laboratory specialists prefer using the 1:1 mix of patient plasma and normal control plasma for initial mixing studies, occasionally using a 3:1 or 4:1 mix (4 parts patient plasma to one part normal control plasma) when a thrombotic clinical picture suggests a lupus anticoagulant but the 1:1 mix corrects. The latest guidelines from the ISTH Standardization Subcommittee also specify 1:1, see Pengo V, Tripodi A, Reber G, et al. Update of the guidelines for lupus anticoagulant detection. J Thromb Haemost 2009; 7: 1737-40.

I consulted with Dr. Larry Brace of Edward Hospital, Naperville, IL about this and he also advocates for the 1:1 mix on the grounds that a lupus anticoagulant-sensitive partial thromboplastin time (PTT) reagent such as PTT–LA is at least as likely to detect lupus anticoagulant as is a 4:1 mix. He also reminded me that lupus anticoagulant testing must always include at least two test systems. Most of us use a lupus anticoagulant sensitive PTT reagent system and a dilute Russell viper venom system (DRVVT), although many continue to use the dilute prothrombin time (PT) assay and the kaolin clotting time. These latter two are usually done manually. Both systems must include a high phospholipid concentration reagent neutralization step for confirmation.

There are many experts who argue for the 4:1 mix, first developed by Dr. Doug Triplett in the 1970s. These include my friend Dan Kaczor of Diagnostica Stago-US, and I have sent him an e-mail asking for his response to your question. Dan received his coagulation lab education from Dr. Triplett.

I posted a Quick Question in June polling our participants on their criteria for mixing study correction. Click here to see the responses, which are inconclusive. It looks like the majority of people use correction to within 5 seconds of the normal control plasma. I prefer correction within 10%, which is virtually identical to the Rosner index, whereas Dr. Dorothy Adcock of Esoterix Coagulation requires correction to within the  upper limit of the normal range. There are no clinical studies that support the efficacy of one over the other, and the Pengo article does not address the argument!

I hope this is helpful, and I’m sure we will collect additional comments. Geo.