The Principles of Clinical Cytogenetics - Extra Materials - Springer

Quality Control and Quality Assurance 99
• The slide temperature
• Wet or dry slide? How much water?
• The angle of the slide during specimen application
• The method of applying the specimen
• The method of drying the slide
• The slide-aging technique
Each of these factors significantly contributes to the success of slide preparation. As theses can be
variable from day to day or between individuals, close observation and documentation of technique
can allow the highest proficiency of these skills.
Banding and Staining
Slide preparation and aging are deciding factors in the lab’s ability to successfully stain a specimen,
by affecting adjustments to solution concentration, the time a slide is left in a staining solution,
and so forth. Careful reagent preparation and documentation of adjustments made to staining procedures
helps laboratories to refine techniques.
The shelf life and storage conditions of banding and staining reagents are important considerations
and should also be documented in a staining log. As reagents arrive in the laboratory, lot
numbers should be recorded and compared with previous lots used. Reagent containers should be
labeled with the reagent name, quantity, concentration, storage requirements, the date received, and
an expiration date. Reagents that require refrigeration should have minimum and maximum permissible
temperatures documented, and these should not be exceeded. Existing supplies of reagents
should be rotated so that they are depleted before new supplies are used.
Although good specimen staining is important for microscope analysis, it is also necessary to
consider the microscope on which a specimen will be analyzed and the staining requirements of the
recording media (photography or electronic image capture). When a laboratory has a variety of microscopes,
each might have a light source, contrast or interference filters, objectives, or other lenses that
produce images with a unique set of visual characteristics. Additional variables are introduced with
the use fluorescence microscopy, such as excitation and barrier filters, and features such as the numerical
aperture of lenses or bulb intensity could be critical. Individual taste will also play an
important factor in identifying a staining intensity that is well suited for microscope analysis.
For either photography or electronic capture of traditional G-banded images, it is important to
identify staining intensities that produce the following:
• Chromosome pale ends that contrast well against background areas
• A wide range in mid-gray intensity
• Dark bands in close proximity appear as distinct bands
It is not unusual to find that a staining protocol might not be well suited for both microscope
analysis and photography/electronic capture.
Comparing the requirements of the individual performing the microscope analysis against the
requirements of the recording media and documenting the ideal conditions in a staining log will help
laboratories gain control of the many variables of a staining procedure.
Specimen Analysis
Any chromosome analysis must begin by identifying specific requirements for the specimen type
being examined. Following this, the basic steps involved are microscope analysis itself (location of
metaphase spreads suitable for analysis, counting chromosomes and determining sex chromosome
complement, and band-pattern analysis), photographic or computerized imaging, preparation of
karyotypes, and documentation and reporting of results. The procedure begins with a protocol that
must be accessible and thoroughly understood by all individuals performing chromosome analysis.