Immunomagnetic separation of Cryptosporidium parvum oocysts using MACS MicroBeads and high gradient separation columns

We evaluated the MACS immunomagnetic separation (IMS) system for concentrat ing Cryptosporidium parvum. Oocysts were first labeled with fluorescein iso thiocyanate (FITC) or rabbit anti-C parvum antibodies, then linked to Micro Beads coated with anti-FITC or anti-rabbit IgG, and separated through a hig h gradient separation column. Results indicated that over 95% of oocysts we re recovered and their fluorescence and infectivity were retained. The pres ence of MicroBeads showed no effect on genomic DNA extraction and subsequen t polymerase chain reaction (PCR)-based analyses, as sensitivity of PCR (10 oocysts) and the band pattern of randomly amplified polymorphic DNA (RAPD) were identical to those using DNAs extracted from normally purified oocyst s. IMS-PCR consistently detected as few as 10 oocysts from 100 mi of apple juice or homogenized milk and IMS-IFA could detect 100 oocysts from 1 g of deer manure, demonstrating the efficiency of IMS in recovering oocysts from environmental and food samples. Our results suggest that the MACS IMS syst em could be used for multiple applications in Cryptosporidium research. (C) 2000 Published by Elsevier Science B.V. All rights reserved.