Erythrocyte transketolase activity coefficient (ETKAC) assay protocol for the assessment of thiamine status.
Annals of the New York Academy of Sciences 2020 ; 1498: 77-84.
DOI : 10.1111/nyas.14547
PubMed ID : 33354793
PMCID : PMC8451777
Vitamin B1 (thiamine) is an essential nutrient that acts as a cofactor for a number of metabolic processes, particularly in energy metabolism. Symptoms of classic thiamine deficiency are recognized as beriberi, although clinical symptoms are nonspecific and recognition of subclinical deficiency is difficult. Therefore, reliable biomarkers of thiamine status are required. Thiamine diphosphate is a cofactor for transketolase, including erythrocyte transketolase (ETK). The ETK activity assay as an indirect, functional marker of thiamine status has been used for over 50 years. The ETK activity assay provides a sensitive and specific biomarker of thiamine status; however, there is a lack of consensus over the cutoffs for deficiency, partly due to a lack of assay harmonization. Here, we provide a step-by-step protocol for the measurement of ETK activity and the calculation of the ETK activity coefficient, including detailed explanations of equipment and chemicals required and guidance for quality control procedures. Harmonization of the protocol will provide the basis for the development of internationally recognized cutoffs for thiamine insufficiency. The establishment of quality control materials and a quality assurance scheme are recommended to provide reliability. This will ensure that the ETK activity assay remains an important method for the assessment of thiamine status.
Vitamin B1 (thiamine) is an essential nutrient required for energy metabolism and the nervous system. Thiamine deficiency can cause infantile beriberi. Populations particularly at risk of thiamine deficiency are breastfed infants of thiamine-deficient mothers in low-income countries, especially those where rice, which contains little thiamine, is the staple food. However, deficiency is associated with a range of non-specific clinical symptoms and can be difficult to diagnose. Evidence also exists to suggest that mild thiamine deficiency may have long-term effects on brain development and gross motor skills.
Biomarkers are compounds we can measure in blood that tell us about a person’s physiology and health. Biomarkers of thiamine status are essential to identify deficiency and improve understanding of the global prevalence of thiamine deficiency and of the links between thiamine and later health outcomes.
An important biomarker of thiamine status is the “erythrocyte transketolase activity coefficient” (ETKAC). ETKAC is a measure of the availability of thiamine available for use in in red blood cells (erythrocytes). The ETKAC assay has been available for over 50 years but few laboratories offer the assay, partly due to the complexity of older methods. Development of the assay for use in 96-well plates with microplate reader resulted in a more cost-effective and efficient technique.
In this manuscript, we provide a step-by-step protocol to perform the ETKAC assay. We include detailed descriptions of equipment and methods and describe the quality control procedures.
This protocol will facilitate harmonisation of the ETKAC assay. It provides a foundation for the establishment of the assay in new laboratories and supports the investigation of outstanding questions in thiamine biology contributing to the ultimate aim of developing strategies to control thiamine deficiency.