Fm. Vanderijke et al., SAMPLE PREPARATION AND IN-SITU HYBRIDIZATION TECHNIQUES FOR AUTOMATEDMOLECULAR CYTOGENETIC ANALYSIS OF WHITE BLOOD-CELLS, Cytometry, 24(2), 1996, pp. 151-157
With the advent of in situ hybridization techniques for the analysis o
f chromosome copy number or structure in interphase cells, the diagnos
tic and prognostic potential of cytogenetics has been augmented consid
erably. In theory, the strategies for detection of cytogenetically abe
rrant cells by in sim hybridization ate simple and straightforward. In
practice, however, they are fallible, because false classification of
hybridization spot number or patterns occurs. When a decision has to
be made on molecular cytogenetic normalcy or abnormalcy of a cell samp
le, the problem of false classification becomes particularly prominent
if the fraction of aberrant cells is relatively small. In such mosaic
situations, often > 200 cells have to be evaluated to reach a statist
ical sound figure. The manual enumeration of in situ hybridization spo
ts in many cells in many patient samples is tedious. Assistance in the
evaluation process by automation of microscope functions and image an
alysis techniques is, therefore, strongly indicated, Next to research
and development of microscope hardware, camera technology, and image a
nalysis, the optimization of the specimen for the (semi)automated micr
oscopic analysis is essential, since factors such as cell density, thi
ckness, and overlap have dramatic influences on the speed and complexi
ty of the analysis process. Here we describe experiments that have led
to a protocol for blood cell specimen that results in microscope prep
arations that are well suited for automated molecular cytogenetic anal
ysis. (C) 1996 Wiley-Liss, Inc.