The Principles of Clinical Cytogenetics - Extra Materials - Springer

116 Steven Gersen and Lotte Downey
Fig. 3. Benchtop drying chamber. (Courtesy of Percival Scientific, Inc.)
As the 3:1 methanol:acetic acid fixative used in cytogenetics laboratories dries, it “pulls” the cell
membrane across the slide or cover slip with it, allowing the chromosomes of mitotic cells to separate.
If this process is viewed with a phase-contrast microscope, the metaphases appear to open much
like a flower blossom. Clearly, the ambient temperature and humidity, as well as airflow over the
cells (and possibly, as suggested by some studies, the barometric pressure) all affect the rate of drying;
therefore, when utilizing in situ processing, controlling these parameters is the only way to
control chromosome spreading (1).
In fact, of greatest importance is not merely controlling conditions, but maintaining them with a
high degree of consistency. With each change in any one parameter, drying and spreading of chromosomes
changes; once the correct combination is achieved, it is of paramount importance that it be
maintained throughout the entire harvest.
There are probably as many solutions to this situation as there are cytogenetics laboratories. Some
have constructed enclosed chambers in which airflow, humidity, and temperature can be varied,
although these are typically prone to failure whenever the air conditioning breaks, because it is easy
to warm the air inside the chamber but extremely difficult to cool it. Some labs have designed climatecontrolled
rooms; these frequently function well, but the drawbacks here are the need to maintain
conditions while properly venting out fixative fumes (an engineering challenge, but certainly possible)
and the potential to expose the technologist to uncomfortable conditions. Such rooms are also
often costly to build.
Recently, several companies have developed self-contained chambers specifically for the purpose
of drying in situ cultures; an example is shown in Fig. 3. Initially developed for the culture of insect
cells (which are grown at room temperature, and so the incubator must be capable of cooling as well