PMID- 11076056
OWN - NLM
STAT- MEDLINE
DCOM- 20010301
LR  - 20190816
IS  - 0009-9163 (Print)
IS  - 0009-9163 (Linking)
VI  - 58
IP  - 4
DP  - 2000 Oct
TI  - Optimization of the fluorescence in situ hybridization (FISH) technique for high 
      detection efficiency of very small proportions of target interphase nuclei.
PG  - 309-18
AB  - Using commercially available fluorochrome-labeled probes specific for chromosomes 
      X, Y, 13, 18, and 21, we optimized the technical protocols for fluorescence in 
      situ hybridization (FISH) so that the highest sensitivity and specificity were 
      achieved. Also, we compared the optical properties of different types of 
      fluorescent labels in an effort to develop the most efficient FISH protocol, 
      including the determination of which types of labels are the easiest to count 
      accurately. The lymphocytes were purified from blood of normal male and female 
      newborns, normal male and female adults, and a trisomy 21 male adult. Male and 
      female lymphocytes were mixed in five different combinations. For each 
      combination, the male lymphocytes either from newborns or from adults were 
      diluted with female lymphocytes in seven different proportions. For each of these 
      35 different cell mixtures, 100,000 nuclei were analyzed and scored in a blind 
      fashion. Among the different fluorochrome-labeled probes, the highest sensitivity 
      and specificity were achieved when SpectrumAqua CEP-Y/SpectrumOrange CEP X probe 
      mixture, SpectrumAqua CEP-18, SpectrumOrange LSI-13, and SpectrumOrange LSI-21 
      were hybridized. The hybridization sensitivity and specificity were higher than 
      99% for the identification of chromosomes X, Y, 13, and 18, and higher than 98% 
      for the detection of trisomy 21. The proportion of false-positive signals was 
      under 0.005% for XY detection and lower than 0.14% for autosome detection. With 
      these high hybridization sensitivities and specificities, the optimized FISH 
      protocol developed in our laboratory has the potential to detect very rare 
      events, e.g., when the proportion of cells being sought is lower than 0.01%. In 
      other words, our protocol allows the specific detection of one male cell sunken 
      among 10,000 female cells.
FAU - Yan, J
AU  - Yan J
AD  - Department of Medical Biology, Universite Laval and Hopital, Saint-Francois 
      d'Assise, CHUQ, Quebec, Canada.
FAU - Guilbault, E
AU  - Guilbault E
FAU - Masse, J
AU  - Masse J
FAU - Bronsard, M
AU  - Bronsard M
FAU - DeGrandpre, P
AU  - DeGrandpre P
FAU - Forest, J C
AU  - Forest JC
FAU - Drouin, R
AU  - Drouin R
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - Denmark
TA  - Clin Genet
JT  - Clinical genetics
JID - 0253664
RN  - 0 (Fluorescent Dyes)
SB  - IM
MH  - Adult
MH  - Cell Nucleus/*ultrastructure
MH  - Chromosomes, Human, Pair 13
MH  - Chromosomes, Human, Pair 18
MH  - Chromosomes, Human, Pair 21
MH  - False Positive Reactions
MH  - Female
MH  - Fluorescent Dyes/pharmacology
MH  - Humans
MH  - In Situ Hybridization, Fluorescence/*methods
MH  - Infant, Newborn
MH  - *Interphase
MH  - Lymphocytes/ultrastructure
MH  - Male
MH  - Sensitivity and Specificity
MH  - X Chromosome
MH  - Y Chromosome
EDAT- 2000/11/15 11:00
MHDA- 2001/03/07 10:01
CRDT- 2000/11/15 11:00
PHST- 2000/11/15 11:00 [pubmed]
PHST- 2001/03/07 10:01 [medline]
PHST- 2000/11/15 11:00 [entrez]
AID - 10.1034/j.1399-0004.2000.580409.x [doi]
PST - ppublish
SO  - Clin Genet. 2000 Oct;58(4):309-18. doi: 10.1034/j.1399-0004.2000.580409.x.