416
T. C. HSU
We know much more about the activities of the interphase cell now
than we did two decades ago. The establishment of the concept of
DNA constancy was a major contribution toward understanding
chromosome synthesis. Thus if a method could be devised to measure
the D N A content of a nucleus without chemical extraction, one should
be able to study the events associated with D N A synthesis during
interphase, which could then be transcribed to chromosome synthesis.
The development of microspectrophotometry fulfilled, at least in part,
this need. With a standard D N A staining procedure, the Feulgen
reaction, the amount of the stain, which represents the amount of DNA
present in a nucleus, can be measured by passing a beam of suitable
monochromatic light through the nucleus. The constancy of D N A
content per cell can best be established by measuring that of individual
sperm cells since they are known to contain a haploid set of chromosomes. Using this value as a basis, the D N A content of somatic cells
can also be recorded (cf. Swift, 1953).
The painstaking measurements made by various workers confirmed the hypothesis that D N A is indeed synthesized during interphase
(Swift, 1950; Walker and Yates, 1952). It has also been shown that
D N A synthesis occurs only during a limited portion of interphase
before the onset of mitosis. In mammalian cells, Richards, Walker and
Deeley (1956) calculated synthesis curves from the frequency distribution of amounts of D N A in interphase cells. (See also Chapter 7.)
A. AUTORADIOGRAPHY
The microspectrophotometric technique has a number of applications but it also has a number of limitations. It cannot tell at exactly
which time of interphase the D N A synthesis is active. It cannot tell
with precision how long it lasts, when it begins and when it ends. It
cannot be applied to R N A studies. Some of these difficulties have been
resolved by autoradiographic studies utilizing tritium-labeled nucleic
acid precursors or analogues.
Utilization of radioactive isotopes has yielded much information in
biology and biochemistry, particularly in elucidating intermediary
metabolism. Popular isotopes are carbon-14, sulfur-35, phosphorus-32,
and nitrogen-15. For chemical studies, these suffice because one can
isolate and purify compounds and record radioactivity with accurate
instruments. However, to attack histological and cytological problems,
one must observe microscopically the tissues, cells or cell components
and at the same time locate the isotope label. If a photographic film
is placed over the tissue sections or cells which bear the label, the
j8-rays emitted by the decay of the isotope expose the film. After the
T. C. HSU
We know much more about the activities of the interphase cell now
than we did two decades ago. The establishment of the concept of
DNA constancy was a major contribution toward understanding
chromosome synthesis. Thus if a method could be devised to measure
the D N A content of a nucleus without chemical extraction, one should
be able to study the events associated with D N A synthesis during
interphase, which could then be transcribed to chromosome synthesis.
The development of microspectrophotometry fulfilled, at least in part,
this need. With a standard D N A staining procedure, the Feulgen
reaction, the amount of the stain, which represents the amount of DNA
present in a nucleus, can be measured by passing a beam of suitable
monochromatic light through the nucleus. The constancy of D N A
content per cell can best be established by measuring that of individual
sperm cells since they are known to contain a haploid set of chromosomes. Using this value as a basis, the D N A content of somatic cells
can also be recorded (cf. Swift, 1953).
The painstaking measurements made by various workers confirmed the hypothesis that D N A is indeed synthesized during interphase
(Swift, 1950; Walker and Yates, 1952). It has also been shown that
D N A synthesis occurs only during a limited portion of interphase
before the onset of mitosis. In mammalian cells, Richards, Walker and
Deeley (1956) calculated synthesis curves from the frequency distribution of amounts of D N A in interphase cells. (See also Chapter 7.)
A. AUTORADIOGRAPHY
The microspectrophotometric technique has a number of applications but it also has a number of limitations. It cannot tell at exactly
which time of interphase the D N A synthesis is active. It cannot tell
with precision how long it lasts, when it begins and when it ends. It
cannot be applied to R N A studies. Some of these difficulties have been
resolved by autoradiographic studies utilizing tritium-labeled nucleic
acid precursors or analogues.
Utilization of radioactive isotopes has yielded much information in
biology and biochemistry, particularly in elucidating intermediary
metabolism. Popular isotopes are carbon-14, sulfur-35, phosphorus-32,
and nitrogen-15. For chemical studies, these suffice because one can
isolate and purify compounds and record radioactivity with accurate
instruments. However, to attack histological and cytological problems,
one must observe microscopically the tissues, cells or cell components
and at the same time locate the isotope label. If a photographic film
is placed over the tissue sections or cells which bear the label, the
j8-rays emitted by the decay of the isotope expose the film. After the