M. Griffiths, H. Vetr, N. B. Binder

Research and Development Department, Technoclone Herstellung von Diagnostika und Arzneimitteln GmbH, Vienna, Austria

Introduction

Background
Severe deficiency in ADAMTS-13 activity levels (< 0.1 IU/ml) is a specific marker for thrombotic thrombocytopenic purpura (TTP). TTP is an acute life threatening disorder with the highest mortality rate within the first 24 h of admission. It would be beneficial to have a quick turnaround of ADAMTS-13 activity reporting. We have developed and validated a simple screening assay which allows the reporting within 1 hour of blood draw.

Materials and Methods

Normal and Thromobocytopenic Purpura (TTP) patient plasma samples were tested in the TECHNOZYM® ADAMTS-13 Activity ELISA and the TECHNOSCREEN® ADAMTS-13 Activity. A cut off value of 0.1 IU/ml was chosen since this reflects the value used by laboratories for classifying a TTP patient. The sample results were classified as either > 0.1IU/mL or <0.1IU/mL in order to calculate the parameters below.

Results

TECHNOSCREEN® ADAMTS-13 Activity Assay

  • Based on flow through technology.
  • Plasma sample is pre-incubated with ADAMTS-13 specific substrate for 20 minutes at room temperature
  • A drop of pre-incubation mixture is added to flow through unit containing a test & control well (see fig 1.& 2)
  • In the test well the immobilized antibody (G7) will bind cleaved substrate only
  • In the control well the immobilized antibody (anti GST) will bind both cleaved and uncleaved substrate
  • Bound substrate is detected using a biotinylated anti GST antibody and Streptavidin–gold conjugate
  • The intensity of the red colour in the test well is proportional to the level of ADAMTS-13 Activity in the plasma sample and is compared against a colour standard card, allowing for semi quantitation of activity levels.
  • The control well should always be dark red in colour, validating the test result
  • Assay is easy to run as all the reagents are added with dropper bottles
  • Total assay time is <30 minutes.

Fig 2. Examples of normal & TTP sample in TECHNOSCREEN® ADAMTS-13 Activity Test

Performance Data

Conclusions

We have demonstrated that this screening test is an excellent tool for determining deficient levels of ADAMTS-13 activity with a sensitivity >96%.

The TECHNOSCREEN ADAMTS-13 Activity is:

  • Designed for testing single patient samples
  • Versatile as can be performed without specialized laboratory equipment
  • Ideal as a cost effective screening tool for TTP

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