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Table 1 Testing for presence of PrPSc in BSE samples subsequently treated with heat and digestion by keratinase

From: Stability of BSE infectivity towards heat treatment even after proteolytic removal of prion protein

1, test (principle)

2, antibody

3, measuring unit

4, test-control

5, non-heated / digested

6, heated / digested

7, PrPSc signal as % of non-heated digested (reduction factor)c

TeSeE (capture)a

 SAF32, Bar224

 OD

 2.288b

 0.020 ± 0.016

 0.009 ± 0.003 [0.012]

 <  < 0.04% (> 2500)

(0.009 ± 0.004) [0.013]

HerdCheck (Seprion ligand binding)

?

OD

3.73

1.93 ± 1.13 [3.06b]

0.07 ± 0.04 [0.11]

 <  < 0.3% (> 333)

(0.06 ± 0.06) [0.12]

CediTect BSE (conformation dependent)

9A2

d/n

170

131.8 ± 48.5 [180.3b]

1.5 ± 0.5 [2.0]

 < 0.2% (> 500)

(1.6 ± 0.1) [1.7]

94B4

d/n

255

141.8 ± 91.4 [233.2b]

2.1 ± 1.3 [3.4]

 < 0.04% (> 2500)

(2.0 ± 1.4) [3.4]

  1. Values in tests 1—3 are presented as averages ± SD of four BSE positive cases. Between parentheses BSE negative cases. Between brackets sum of average plus maximum SD values.
  2. Absence of PrPSc in heated and protease digested bovine brain homogenates. Differently treated homogenates from confirmed BSE positive cattle (n = 4, two clinical cases, two cases found at healthy slaughter) were subjected to three different immunochemical tests. Also four similarly treated homogenates from confirmed BSE negative brains were included. Values in columns 5 and 6 are presented as averages ± SD of four cases; between parentheses values found in the confirmed BSE negative cases. Between brackets sum of average value plus maximum SD value. Only the IDEXX and CediTect tests appeared able to recognize PrPSc in non-heated and KE-digested samples. The BSE positive reference controls—column 4—in the TeSeE and CediTect tests have been digested with PK exactly according to the manufacturer’s instructions and were performed on regular BSE positive samples from routine testing. The IDEXX test recognizes intact PrPSc as well as PrPres. The CediTect assay was applied with antibodies 9A2 and 94B4 which are specific respectively for the N-terminal and C-terminal region of PrPres. Tissue amounts per tested well were for TeSeE, IDEXX and CediTect test respectively 3, 0.6 and 0.5 mg wet tissue weight per well.
  3. aThe Biorad TeSeE ELISA does not recognize PrPres in the non-heated KE digests (column 5) since the PrP octarepeat needed for recognition by the capture antibody SAF34 is fully removed during digestion with the keratinase where the conditions differ from those used for the test control. Therefore, for this ELISA the value of the positive test control in column 4 was used for calculation of the percentage residual PrPSc in column 7 since the digested non-heated cases do not bind to the catching antibody where the correct conditions for retaining the SAF34 epitope could not be applied in our digestion system with keratinase.
  4. bThe 100% PrPSc values of the samples in the different tests before heat treatment were used for calculating the fraction of remaining PrPSc signals of column 7.
  5. cCalculations for column 7: value of BSE positives (column 6, average value plus maximum SD) – value of BSE negatives (column 6 between parenteses, average value plus maximum SD) / 100% PrPSc-values mentioned in footnote b.
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Veterinary Research

ISSN: 1297-9716