PMID- 16793153
OWN - NLM
STAT- MEDLINE
DCOM- 20061212
LR  - 20131121
IS  - 0167-7012 (Print)
IS  - 0167-7012 (Linking)
VI  - 67
IP  - 2
DP  - 2006 Nov
TI  - Direct counting of Cryptosporidium parvum oocysts using fluorescence in situ 
      hybridization on a membrane filter.
PG  - 373-80
AB  - This report describes the development of a direct and rapid detection method for 
      the pathogenic protozoan, Cryptosporidium parvum, from environmental water 
      samples using fluorescence in situ hybridization (FISH) on a membrane filter. The 
      hydrophilic polytetrafluoroethylene (PTFE) membrane filter with FISH-stained 
      oocysts yielded the highest signal to noise (S/N) ratio of the different membrane 
      filters tested. PTFE membranes retained 98.8+/-0.4% of the concentrated oocysts 
      after washing, simultaneous permeabilization and fixation with a hot ethanol 
      solution, and hybridization with a fluorescently labeled oligonucleotide probe. 
      This procedure eliminates subsequent time-consuming recovery steps that often 
      result in a loss of the actual oocysts in a given environmental water sample. 
      Furthermore, C. parvum was successfully distinguished from Cryptosporidium muris 
      and other species in environmental water samples with the addition of formamide 
      into the hybridization solution. In tap water samples, the S/N ratio was 
      heightened by washing the membrane filter prior to FISH with a 1 M HCl solution 
      in order to reduce the large amounts of impurities and background fluorescence 
      from the non-specific adsorption of the fluorescently labeled oligonucleotide 
      probe.
FAU - Taguchi, Tomoyuki
AU  - Taguchi T
AD  - Department of Biotechnology, Tokyo University of Agriculture and Technology, 
      2-24-16 Naka-cho, Koganei, Tokyo 184-8588, Japan.
FAU - Shinozaki, Youhei
AU  - Shinozaki Y
FAU - Takeyama, Haruko
AU  - Takeyama H
FAU - Haraguchi, Satoshi
AU  - Haraguchi S
FAU - Yoshino, Masato
AU  - Yoshino M
FAU - Kaneko, Masao
AU  - Kaneko M
FAU - Ishimori, Yoshio
AU  - Ishimori Y
FAU - Matsunaga, Tadashi
AU  - Matsunaga T
LA  - eng
PT  - Journal Article
DEP - 20060621
PL  - Netherlands
TA  - J Microbiol Methods
JT  - Journal of microbiological methods
JID - 8306883
RN  - 0 (Carbocyanines)
RN  - 0 (DNA Probes)
RN  - 0 (Fluorescent Dyes)
RN  - 0 (Membranes, Artificial)
RN  - 0 (cyanine dye 3)
RN  - 059QF0KO0R (Water)
SB  - IM
MH  - Animals
MH  - Carbocyanines/chemistry
MH  - Cryptosporidium parvum/genetics/*isolation & purification
MH  - DNA Probes/genetics
MH  - Fluorescent Dyes/chemistry
MH  - Humans
MH  - In Situ Hybridization, Fluorescence/*methods
MH  - Membranes, Artificial
MH  - Microscopy, Fluorescence
MH  - Microscopy, Interference
MH  - Oocysts
MH  - Water/*parasitology
EDAT- 2006/06/24 09:00
MHDA- 2006/12/13 09:00
CRDT- 2006/06/24 09:00
PHST- 2006/01/17 00:00 [received]
PHST- 2006/03/30 00:00 [revised]
PHST- 2006/04/18 00:00 [accepted]
PHST- 2006/06/24 09:00 [pubmed]
PHST- 2006/12/13 09:00 [medline]
PHST- 2006/06/24 09:00 [entrez]
AID - S0167-7012(06)00120-5 [pii]
AID - 10.1016/j.mimet.2006.04.017 [doi]
PST - ppublish
SO  - J Microbiol Methods. 2006 Nov;67(2):373-80. doi: 10.1016/j.mimet.2006.04.017. 
      Epub 2006 Jun 21.