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Evaluation of co‐oximetry for the measurement of methemoglobin in rainbow trout ( O ncorhynchus mykiss ) and values in 3 salmonid species
Author(s) -
Saunders Janet,
Speare David J.,
McConkey Sandra
Publication year - 2012
Publication title -
veterinary clinical pathology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.537
H-Index - 51
eISSN - 1939-165X
pISSN - 0275-6382
DOI - 10.1111/j.1939-165x.2012.00466.x
Subject(s) - methemoglobin , rainbow trout , hemoglobin , chemistry , chromatography , biochemistry , fish <actinopterygii> , biology , fishery
Background Methemoglobin (met H b) is oxidized hemoglobin that cannot reversibly bind oxygen, and concentrations in healthy fish have been reported to be 0.6–24.8% compared with 0–3% in healthy mammals. In fish, met H b has been measured using spectrophotometric methods using potassium cyanide ( KCN ), but not using co‐oximetry, which is the preferred method for human samples. Objectives The aims of this study were to evaluate co‐oximetry as a method for measuring metHb in O ncorhynchus mykiss , compare co‐oximetry with a KCN spectrophotometric method, and establish reference values for metHb concentrations as measured using co‐oximetry in O mykiss, Salmo salar, and Salvelinus fontinalis . Methods Blood samples from healthy female O mykiss, female S salar , and female and male S fontinalis were prepared by separation and washing of erythrocytes in Tris/NaCl/ EDTA buffer followed by lysis in Tris/ EDTA buffer. MetHb concentrations were measured using an IL ‐682 co‐oximeter. Moderate and high metHb concentrations were produced in vitro using NaNO 2 . Results At low concentrations of methemoglobin, CV s for intraday precision were 10.3% and 53.9% using co‐oximetry and the KCN spectrophotometric method, respectively. The CV for interday precision using co‐oximetry was 11.9%. Met H b concentrations were stable in whole blood stored at 4°C for 7 days. Met H b concentrations were linear up to 58.2% ( r = .99) using co‐oximetry and 27.5% ( r = .94) using the KCN method. The lower limit of detection for met H b was 0.02 g/dL using co‐oximetry. Reference values for met H b concentrations using co‐oximetry in O mykiss, S salar, and S fontinalis ( n = 40 of each species) were 0.6–1.8%, 1.1–1.9%, and 1.1–4.0%, respectively. Conclusions Co‐oximetry can be used to measure methemoglobin in blood from fish, in particular in O mykiss , and is better than the KCN spectrophotometric method. Reference values for methemoglobin concentrations in O mykiss , S salar , and S fontinalis are similar to those in mammals.