Method for Detection and Quantitation of Fathead Minnow Vitellogenin (Vtg) by Liquid Chromatography and Matrix Assisted Laser Desorption/ Ionization Mass Spectrometry

Journal Article · Fri Mar 11 04:00:00 EST 2005 · Aquatic Toxicology

DOI:https://doi.org/10.1016/j.aquatox.2005.03.015· OSTI ID:859978

Wunschel, David S; Schultz, Irv R; Skillman, Ann D; Wahl, Karen L

Vitellogenin (Vtg) is a well recognized biomarker of estrogen exposure in many species, particularly fish. This large protein shares a high degree of sequence homology across a large number of species. Quantitative measurement is currently done using antibody-based assays. These assays frequently require purification of Vtg and antibody production from each species because there is poor cross reactivity between antibodies for different fish. Therefore, complementary methods of measuring Vtg are desirable. Mass spectrometric (MS) analysis coupled to database searching offers the promise of a general method for protein identification. In this study we used the well characterized Vtg from rainbow trout (O. mykiss) to evaluate the analytical parameters for matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) analysis of the intact and tryptic digested protein. An analytical scale HPLC separation combined with MALDI-MS was used to measure and confirm the identity of Vtg from the plasma of an important species for regulatory agencies, fathead minnow (Pimephales promelas). The small volume requirement of this method (< 10 uL) was found to be compatible with the plasma volume obtained from a few minnows. A semi quantitative measurement of Vtg from minnows exposed to estradiol was achieved, which was similar to previously obtained ELISA data.

• Research Organization: Pacific Northwest National Laboratory (PNNL), Richland, WA (US)
• Sponsoring Organization: USDOE
• DOE Contract Number: AC05-76RL01830
• OSTI ID: 859978
• Report Number(s): PNWD-SA-6963
• Journal Information: Aquatic Toxicology, Journal Name: Aquatic Toxicology Vol. 73; ISSN AQTODG; ISSN 0166-445X
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
60 APPLIED LIFE SCIENCES
ANTIBODIES
CHROMATOGRAPHY
DESORPTION
DETECTION
ENZYME IMMUNOASSAY
ESTRADIOL
ESTROGENS
FATHEAD MINNOW
HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY
IONIZATION
LASERS
MASS SPECTROSCOPY
PROTEINS
PURIFICATION
TROUT
Vitellogenin
Ethynyl Estradiol
HPLC
MALDI-MS
Peptide Mass Fingerprinting