title: Measurement of the noncomplexed free fraction of tissue inhibitor of metalloproteinases 1 in plasma by immunoassay.
abstract: BACKGROUND
  We previously found differences in total concentrations of tissue inhibitor of metalloproteinases 1 (TIMP-1) in plasma from donors and cancer patients. Because TIMP-1 can exist in more than one molecular form, a new immunoassay to specifically detect free TIMP-1 was developed and concentrations were determined in plasma from healthy donors and colorectal cancer (CRC) patients.
  
  
  METHODS
  We established and validated an immunoassay for the specific measurement of free TIMP-1 that uses a polyclonal anti-TIMP-1 antibody for capture and a monoclonal anti-TIMP-1 antibody that binds only free TIMP-1 for detection of antigen. Plasma samples from healthy donors and CRC patients were assayed for free TIMP-1. Total TIMP-1 was measured by our previously published assay.
  
  
  RESULTS
  The mean (SD) concentrations of free TIMP-1 were similar in citrate [55.5 (11.5) microg/L] and EDTA plasma [58.9 (13.3) microg/L] from 76 donors (r(2) = 0.82). In 154 donors, the ratio of free TIMP-1 [mean (SD), 64.5 (18.0) microg/L] to total TIMP-1 [83.8 (19.8) microg/L] in EDTA plasma was 0.77. Plasma concentrations of free and total TIMP-1 correlated significantly to age (free, r(2) = 0.19; total, r(2) = 0.27; P <0.0001), increasing 50% over an age span of 45 years. Free and total TIMP-1 were significantly increased in CRC patients (P <0.0001), whereas the ratio of free to total TIMP-1 (mean, 0.58) was significantly lower than in donors.
  
  
  CONCLUSIONS
  Most of the TIMP-1 in donor plasma is present in its free form, and free TIMP-1 increases with age. Free and total TIMP-1 are increased in CRC patient plasma, but the ratio of free to total TIMP-1 is significantly lower in these patients than in donors.
authors.affiliation: The Finsen Laboratory, Rigshopitalet, Strandboulevarden 49, 2100 Copenhagen, Denmark. mads@finsenlab.dk
authors.forename: Mads N
authors.initials: MN
authors.lastname: Holten-Andersen
authors.forename: Ib Jarle
authors.initials: IJ
authors.lastname: Christensen
authors.forename: Hans Jørgen
authors.initials: HJ
authors.lastname: Nielsen
authors.forename: Hans
authors.initials: H
authors.lastname: Lilja
authors.forename: Gillian
authors.initials: G
authors.lastname: Murphy
authors.forename: Vibeke
authors.initials: V
authors.lastname: Jensen
authors.forename: Nils
authors.initials: N
authors.lastname: Brünner
authors.forename: Timo
authors.initials: T
authors.lastname: Piironen
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000925:Anticoagulants; D019343:Citric Acid; D015179:Colorectal Neoplasms / Q000097:blood; D004492:Edetic Acid; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D008666:Metalloendopeptidases / Q000378:metabolism; D008875:Middle Aged; D012016:Reference Values; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000097:blood* / Q000378:metabolism
chemical_list: D000925:Anticoagulants; D019715:Tissue Inhibitor of Metalloproteinase-1; D019343:Citric Acid; D004492:Edetic Acid; D008666:Metalloendopeptidases