IMPROVEMENT OF PRPSC-DETECTION IN MOUSE SPLEEN EARLY AT THE PRECLINICAL STAGE OF SCRAPIE WITH COLLAGENASE-COMPLETED TISSUE HOMOGENIZATION AND SARKOSYL-NACL EXTRACTION OF PRPSC

Scrapie in sheep has recently become again a target of control measure s and eradication programs. Crucial for the effectiveness of these mea sures is the detection of infected sheep during the long and potential ly hazardous incubation period. However, routine-diagnosis is mostly l imited to clinical examinations when disease: becomes apparent, and to postmortem investigations. Through the detection of the scrapie-speci fic isoform of the pi-ion protein (PrPSc) by Western blot in the splee n and lymph nodes from scrapie-infected mice and sheep, we have shown previously that diagnosis during the preclinical stage is possible. We introduce here an improved method for the diagnosis of mouse scrapie shortly after infection. Through a homogenization procedure that inclu des a collagenase digestion step, and through extraction and salting-o ut of PrPSc by Sarkosyl and NaCl, respectively, we were able to detect PrPSc in spleen tissue of intraperitoneally infected mice seven days postinfection. Moreover, the new protocol makes sample-handling easier and reduces the hands-on time. We also successfully enriched PrPSc fr om spleen tissue through immobilized metal affinity chromatography (IM AC); however, for the diagnosis at the earliest stage of infection, ex traction of PrPSc by Sarkosyl and NaCl was more effective.