Figure 4

WB images from primary passage samples. A and B Nine representative recipients, homozygous for alanine (codon 136) detected by Sha31 (A), or P4 (B). (Lane 1, 63/11; lane 2, 3/11; lane 3, 457/11; lane 4 26/12; lane 5 112/12; lane 6, 182/12; lane 7, 3/13; lane 8, 98/11; lane 9, 455/11; AB + , donor bovine L-BSE; B + , bovine C-BSE; S + , ovine Scrapie; BS + , experimental ovine CBSE; S-, negative sheep; M, molecular mass markers. A 1 min exposure, B 10 min exposure). C and D Eight representative recipients homozygous or heterozygous for valine (codon 136), detected by Sha31 (C) or P4 (D), (Lane 1, case 1591/10; lane 2, case 58/11; lane 3, 140/11; lane 4, 267/11; lane 5, 456/11; lane 6, 113/12; lane 7, 4/12; lane 8, 167/12; markers and controls as for A and B. C 1 min exposure, D 10 min exposure). There is low molecular mass migration of the unglycosylated band (arrow), similar to that of the donor bovine L-BSE (AB+), for all but one of the ovine recipients (VRQ/VRQ Lane 8 C, D) regardless of genotype. Similar di-;mono-glycosylated band ratios are also seen in all cases when detected by mAb Sha31 (see Additional file 4). All recipient samples, irrespective of genotype, are also detected with mAb P4 (B, D). In contrast the donor L-BSE case (AB+) is not. Following extraction with a Proteinase K digestion step, may contain a mixture of varying molecular mass fragments, partly due to multiple cleavage sites and variability in resistance of PrP^Sc to the concentration of the enzyme. The two additional lower bands observed in these profiles are regularly observed in diagnostic samples processed in this way. For diagnostic analysis they are disregarded. Only the standard three bands pertaining to the di, mono and un-glycosylated forms of PrP^Sc are considered relevant.