Homocysteine, total


What is measured?

Total homocysteine (tHcy), which includes homocysteine-cysteine mixed disulphide (Hcy-Cys), free homocysteine (fHcy) and bound homocysteine (bHcy), after quantitative reductive cleavage of all disulphide bonds.
Method(s): GC-MS/MS (1), LC-MS/MS (2).

About homocysteine

Homocysteine is formed from S-adenosylhomocysteine during S-adenosylmethionine-dependent transmethylation, and is salvaged back to methionine in a reaction catalyzed by the enzyme methionine synthase. This enzyme requires methylcobalamin as co-factor and methyltetrahydrofolate as co-substrate, which explains why tHcy increases in folate- or cobalamin-deficient subjects.
More than 100 diseases or conditions are associated with elevated tHcy, and few if any biomarker has been linked to so many pathologies. The causal involvement of homocysteine has often been questioned since lowering of tHcy with B-vitamins did not result in diseases prevention; five diseases can at least in part be prevented by such intervention. These are neural tube defects, impaired childhood cognition, macular degeneration, primary stroke, and cognitive impairment in the elderly (3).


Assessment of folate and cobalamin status; risk factor for cardiovascular disease. Total homocysteine is the preferred metabolic marker of cobalamin status in infants (4).

Specimen, collection and processing

Patient/subject: Prandial status affects concentration, which increases slightly after a protein rich meal..
Matrix: Serum or plasma.
Volume: Minimum volume is 50 µL, but 200 µL is optimal and allows reanalysis.
Preparation and stability: Homocysteine is released from the blood cells, and the resulting artificial increase is inhibited at low temperature. The plasma/serum fraction must be separated from the blood cells, preferentially within 30-60 minutes. After such separation, total homocysteine is stable.


Frozen, on dry ice. (for general instruction on transportation, click here.)

Reported values, interpretation

Reported values: <15 µmol/L. The upper reference limit is lower in in infants (< 6.5 µmol/L) and in pregnant women. Concentration increases in renal failure.
Intraclass correlation coefficient (ICC): 0.72.


1. Midttun, Ø., McCann, A., Aarseth, O., Krokeide, M., Kvalheim, G., Meyer, K., and Ueland, P.M. (2016). Combined measurement of 6 fat-soluble vitamins and 26 water-soluble functional vitamin markers and amino acids in 50 μL of serum or plasma by high-throughput mass spectrometry. Anal Chem 88, 10427-436.
2. Midttun, O., Kvalheim, G., and Ueland, P.M. (2013). High-throughput, low-volume, multianalyte quantification of plasma metabolites related to one-carbon metabolism using HPLC-MS/MS. Anal Bioanal Chem 405, 2009-017.
3. Smith, A. D., & Refsum, H. (2021). Homocysteine – from disease biomarker to disease prevention. J Intern Med, in press.
4. Bjorke-Monsen, A.L., Torsvik, I., Saetran, H., Markestad, T., and Ueland, P.M. (2008). Common metabolic profile in infants indicating impaired cobalamin status responds to cobalamin supplementation. Pediatrics 122, 83-91.

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