Analytical and Bioanalytical Chemistry

, Volume 398, Issue 2, pp 963–972

GC/MS detection of central nervous tissue as specified BSE risk material in meat products and meat and bone meals: thermal stability of markers in comparison with immunochemistry and RT-PCR

  • Ernst Lücker
  • Wolfgang Biedermann
  • Thomas Alter
  • Andreas Hensel
Original Paper

Abstract

Methods for the detection of central nervous tissue (CNT) are urgently needed in food control as a means for controlling strict adherence to both food labeling and banning of specified BSE risk material. Here, we report data on heat stability of the CNT markers neuron-specific enolase (NSE) in western blotting, glial fibrillary acidic protein (GFAP) in an enzyme linked immunoassay, mRNAGFAP in a real-time PCR assay, and several fatty acids (C22:6, C24:0-OH, C24:1ω9/ω7, C24:1ω9-OH/ω7-OH, and C24:0) in gas chromatography mass spectrometry (GC/MS). The sample matrix, a standard material of emulsion-type sausage with varied contents of CNT (brain), was heat-treated in three studies: (1) routine meat technological heat treatment with low (85 °C, 30 min), medium (115 °C, 30 min), and high (133 °C, 30 min, 3 bar) heating of 72 anonymous samples from a blind trial; (2) heat treatment under experimental conditions (100, 110, …, 200 °C, 45 min); and (3) fractionized heating of central nervous system (up to three times) under moderate routine technological conditions (85, 100, and 115 °C, 30 min). The markers of the immunochemical methods showed a low GFAP or very low NSE temperature stability at medium and high temperature conditions. The real-time PCR assay gave inconsistent, non-quantitative results, which indicated an uncontrollable matrix effect. The relevant GC/MS markers (C24:0-OH, C24:1ω9/ω7, and C24:1ω9-OH/ω7-OH) proved to be extremely stable. Neither meat and bone meal conditions (133 °C) nor experimental heating (up to and above 140 °C) showed any reduction of GC/MS CNT quantification. On the contrary, a slight but significant increase was noted over a certain temperature range (120–140 °C) for most fatty acids, possibly due to an improved extractability of the fatty acids. We conclude that a quantitative approach is highly unreliable when using immunochemical methods; moreover, these methods might be basically prone to false-negative results depending on heat treatment and matrix composition. Therefore, antibodies with higher affinity to heat-treated CNT marker epitopes are needed. Relevant amounts of CNT (≥0.5%) in low- and medium-heated products would still be reliably detectable by the GFAP ELISA, which justifies its use as a screening method in official food control. The results obtained by the real-time PCR assay were contradictory to recently published data, indicating a need for further protocol optimization and collaborative trials. Up to date, the analytical approach using GC/MS is the only valid procedure as pertaining to heat stability and quantitative analysis; consequently, it should be recommended as the reference procedure in official food control for CNT detection in heat-treated meat products.

Figure

The introduction of central nervous tissue from bovines into the food chain probably caused a new variant of Creutzfeldt-Jacob disease in humans. Analytical control of meat products by immunochemical CNT detection can be hindered by so far unknown severe heat induced losses. In contrast the CNT-specific fatty acids detected by GC/MS turned out to be remarkably stable up to temperatures of 160 °C

Keywords

Bovine spongiform encephalopathy Central nervous tissue Specified risk material GC/MS ELISA Western blot RT-PCR Fatty acids NSE GFAP 

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Ernst Lücker
    • 1
  • Wolfgang Biedermann
    • 1
  • Thomas Alter
    • 2
  • Andreas Hensel
    • 3
  1. 1.Institut für LebensmittelhygieneUniversität LeipzigLeipzigGermany
  2. 2.Institut für LebensmittelhygieneFreie Universität BerlinBerlinGermany
  3. 3.Bundesinstitut für RisikobewertungBerlinGermany

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