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AN IMPROVED APPARATUS FOR QUANTITATIVE CYTOCHEMICAL MODEL STUDIES AND ITS USE IN AN EXPERIMENTAL TEST OF THE TWO-WAVELENGTH METHOD

P. VAN DUIJN 1, E. M. DEN TONKELAAR 1, and M. J. HARDONK 1

1 Laboratory for Pathology, Biochemical Section, University of Leiden, The Netherlands

The construction of an improved apparatus for quantitative studies on cytochemically-stained membranes is described. The membrane-holder of this apparatus was constructed in such a way that the area of the film whose extinction is measured coincides exactly with the area punched out for weight determination and chemical analyses. In addition, provisions were made to assure an uniform response of the photomultiplier to light rays traversing different parts of the field. Using this system, an experimental test of the so called two-wavelength method was carried out.

The spectra of Feulgen-stained deoxyribonucleic acid cellulose films or periodic acid Schiff-stained cellulose films were measured. The stained membranes were punched out, after which the resulting discs were cut, folded, and brought back to their original position in the holder between two unstained cellulose membranes. In this way the number of chromophores in the measured area was kept constant although their macroscopic distribution had changed considerably.

From the spectra of the folded stained discs the extinction of the same material in unfolded form could be calculated by applying the two-wavelength method of Patau and Ornstein. The two-wavelength method was found to correct well, but some residual error remained even in this macrosystem, due to the relatively strong effect of small measuring errors on the end results.

Submitted on November 29, 1961


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