Development of an updated assay for prekallikrein activator in albumin and immunoglobulin therapeutics

BACKGROUND: Prekallikrein activator (PKA) is a contaminating enzyme found in therapeutic albumin and immunoglobulin products. The level is commonly measured using methods such as that defined by the European Pharmacopoeia (Ph Eur) with traceability to the WHO International Standard for PKA. This method generally works well, but problems are sometimes observed. MATERIALS AND METHODS: A simplified one‐step method has been developed to replace the existing Ph Eur two‐step method which consists of kallikrein generation followed by kallikrein measurement using a chromogenic substrate. Analysis of data from the one‐stage method is simplified by the use of a dedicated online app. RESULTS: The one‐stage method was validated against the current Ph Eur method using batches of albumin and immunoglobulins. Problem batches of immunoglobulins were investigated using the one‐stage method. Improved methodology using true initial rate determinations and use of acid‐treated prekallikrein substrate (PKS) helped understand and reduce artefactual results. CONCLUSIONS: The one‐stage method and associated app streamline real‐time determination of PKA and promote good principles of enzyme assays to limit substrate depletion, while also conserving expensive PKS. Blanking steps and reproducibility are simplified.