This series is going to be a bit different. The top menu gives some brief guidance about the many different techniques that are available. We already have a series following the determination of the development of an uncertainty budget for the simple aPTT. We are comparing and contrasting multiple different methods for calculating the measurement uncertainty and deciding which method is fit for our purpose. In this series we will do the same but for the slightly more complex Factor VIII assay. It is hoped that by following this process we will have a system that we can apply to all assays of coagulation factors

Activity vs concentration v amount

Firstly we will differentiate between measuring the amount of protein (antigenic level) against the activity. It is most common in diagnostic laboratories to test for the activity of Factor VIII. For some reason the definition of the measurand for this factor has always been implemented – we still see FVIII:C (Factor VIII coagulant activity) on request forms to differentiate between other measures of Factor VIII. I have never quite understood why our clinical colleagues have used this approach for only Factor VIII.


Most of the time Factor VIII assays are now performed on automated coagulometers, although rarely they may be done manually. Commonly, it is reserved for teaching purposes. Definition of what analyser is used is useful here. Also the definition of whether it is an optical or mechanical end point detection system that is employed in the assay. There are differences with each technique that we won’t go into here, but it is relevant for our measurand definition.

Similarly, the actual method that is used is very, very important when considering Factor VIII assays. Generally speaking we can detect Factor VIII activity by:

  • Clot based one stage
  • Two stage or
  • Chromogenic

The distinctions between each assay are for another time but it is incredibly important to define which assay method is used as there are significant differences in these assays in specific clinical scenarios. Knowing which assay is performed can inform our clinicians and patients about the meaning of the results produced more effectively.

APTT reagent

There are many different aPTT reagents available, all with different formulations. We discussed this is the aPTT measurand article. It is important to differentiate between each as centres will use different aPTT reagents. As such we should clearly define which reagent is used at the point of measurand definition.


Our Factor VIII activity assays irrespective of whether they are one stage, chromogenic or two stage are always calibrated so this is included in the measurand definition.

Monitoring or diagnosis

The Factor VIII assay is used for two purposes. Diagnosis or determination of a patients Factor VIII level, or monitoring to determine the therapeutic level of a factor replacement therapy. We do not distinguish between the two roles for the Factor VIII assay in the definition o the measurand.

If monitoring, what concentrate/therapy?

A well known hurdle in Factor VIII assays is that there is variability between some of the treatments available (particularly B domain depleted products) and the result that is given for the Factor VIII assay. As such product specific calibrators are used in some cases. The discussion about whether we should be using product specific calibrators for all assays is well beyond the scope of this post, however, it is a topic that is becoming increasing common with the introduction of long acting factor concentrates. We will mention it in the modelling phase. For now, it is sufficient in our measurand definition to define a specific product in the assay if it is the result of a significant assay modification such as different calibrator.




Factor VIII assays can be affected by many other variables that themselves are not part of what is being measured. All factor assays (particularly intrinsic based but also to a lesser extent extrinsic based) can potentially be affected by lupus anticoagulant interference. There are two approaches to performing a factor assay, them being a single point assay where a single measurement is made and the result of that measurement is reported and the more commonly performed, and the recommended approach of performing a series of measurements at different sample dilutions and then averaging the results across those dilutions. Traditionally these are at 1:10, 1:20 and 1:40 dilutions. The purpose here is to see if there is an effect of diluting patient plasma on the calculated result of the factor assay after taking into account the dilution of the patient factor mathematically. This is what is commonly referred to in haemostasis laboratories as factor parallelism. 

That concludes our measurand definition, and the first stage of uncertainty determination for the Factor VIII assay. 

We will move onto modelling next, for a discussion which will allow us to discuss many of the themes that will be applicable to a large subset of our assays in haemostasis. It is fair to say that most of our factor assays can be approached in the same manner as we are using here, so a lot of work is being covered by the initial work we are doing here.