Adaptation of the Diazyme Direct Enzymatic HbA1c Assay for a microplate reader at room temperature

Clin Chem Lab Med. 2011 Jul;49(7):1221-3. doi: 10.1515/CCLM.2011.186. Epub 2011 May 17.


Background: Measurement of hemoglobin A(1c) (HbA(1c)) is a key diagnostic criterion and a key parameter for the follow-up of the treatment of diabetes mellitus. Typically, immunochemical assays of HbA(1c) are performed in clinical chemistry analyzers. In this study, we applied the HbA(1c) assay on a microplate reader at room temperature.

Methods: HbA(1c) samples were measured using the Direct Enzymatic HbA(1c) Assay from Diazyme Laboratories (Poway, CA, USA) using a Plate Chameleon Microplate Reader (Hidex Co., Turku, Finland) according to the manufacturer's protocol and a modification of the method to room temperature. The Tosoh G7 HPLC method for HbA(1c) (Tosoh Co., Tokyo, Japan) was used as a comparative method.

Results: There was good correlation of HbA(1c) results when the assay was performed at room temperature (+22°C) compared with that at +37°C (r=0.987). The modified method was linear over the HbA(1c) range 4%-14%. Analysis of HbA(1c) results from 50 blood samples by the modified method showed good agreement with the HPLC method (r=0.990).

Conclusions: The modified Diazyme Direct Enzymatic HbA(1c) Assay™ appears to work as good at +22°C as that performed according to manufacturer's protocol at +37°C.

MeSH terms

  • Blood Chemical Analysis / methods*
  • Diabetes Mellitus / blood
  • Enzyme Assays
  • Glycated Hemoglobin / analysis*
  • Humans
  • Immunoassay
  • Peroxidase / metabolism
  • Temperature*


  • Glycated Hemoglobin A
  • hemoglobin A1c protein, human
  • Peroxidase