Skip to main content

Testing for Hyperhomocysteinemia in Subjects with a History of Thromboembolic Events Using HPLC Technique

  • Protocol
  • First Online:
Haemostasis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 992))

  • 5948 Accesses

Abstract

An elevated homocysteine level is a well-known thrombophilic risk factor. Determination of total plasma homocysteine therefore is an integrated part of the diagnostic setting after thromboembolic events; about 5–7% of the population do have elevated homocysteine levels. Some forms of hyperhomocysteinemia are treatable; thus a standardized and reliable diagnostic setting has to be at hand. HPLC analysis is widely available in routine diagnostic laboratories. We use the fluorogenic reagent SBD-F to derivate with plasma homocysteine after release of the amino acid from homo- and heterodimers and protein bond using TBP. Separation is performed using a c18 reverse-phase column with aqua and acetonitrile as solvent. Due to continuous release of homocysteine from blood cells centrifugation and separation of plasma within 30 min after venous puncture are crucial for reproducible results.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. D’ Angelo A, Coppola A, Madonna P, Fermo I, Pagano A, Mazzola G, Galli L, Cerbone AM (2000) The role of vitamin B12 in fasting hyperhomocysteinemia and its interaction with the homozygous C677T mutation of the methylenetetrahydrofolate reductase (MTHFR) gene. A case-control study of patients with early-onset thrombotic events. Thromb Haemost 83:563–570

    Google Scholar 

  2. Gaustadnes M, Rudiger N, Rasmussen K, Ingerslev J (2000) Intermediate and severe hyperhomocysteinemia with thrombosis: a study of genetic determinants. Thromb Haemost 83:554–558

    PubMed  CAS  Google Scholar 

  3. Desouza C, Keebler M, McNamara DB, Fonseca V (2002) Drugs affecting homocysteine metabolism: impact on cardiovascular risk. Drugs 62:605–616

    Article  PubMed  CAS  Google Scholar 

  4. Chen P, Poddar R, Tipa EV, Dibello PM, Moravec CD, Robinson K, Green R, Kruger WD, Garrow TA, Jacobsen DW (1999) Homocysteine metabolism in cardiovascular cells and tissues: implications for hyperhomocysteinemia and cardiovascular disease. Adv Enzyme Regul 39:93–109

    Article  PubMed  CAS  Google Scholar 

  5. McCully KS (1996) Homocysteine and vascular disease. Nat Med 2:386–389

    Article  PubMed  CAS  Google Scholar 

  6. Thirup P, Ekelund S (1999) Day-to-day, postprandial, and orthostatic variation of total plasma homocysteine. Clin Chem 45:1280–1283

    PubMed  CAS  Google Scholar 

  7. Jacobsen DW (1998) Homocysteine and vitamins in cardiovascular disease. Clin Chem 44:1833–1843

    PubMed  CAS  Google Scholar 

  8. Araki A, Sako Y (1987) Determination of free and total homocysteine in human plasma by high-performance liquid chromatography with fluorescence detection. J Chromatogr 422: 43–52

    Article  PubMed  CAS  Google Scholar 

  9. Garcia AJ, Apitz-Castro R (2002) Plasma total homocysteine quantification: an improvement of the classical high-performance liquid chromatographic method with fluorescence detection of the thiol-SBD derivatives. J Chromatogr B Analyt Technol Biomed Life Sci 779:359–363

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2013 Humana Press

About this protocol

Cite this protocol

Denecke, J. (2013). Testing for Hyperhomocysteinemia in Subjects with a History of Thromboembolic Events Using HPLC Technique. In: Monagle, P. (eds) Haemostasis. Methods in Molecular Biology, vol 992. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-339-8_31

Download citation

  • DOI: https://doi.org/10.1007/978-1-62703-339-8_31

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-338-1

  • Online ISBN: 978-1-62703-339-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics