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Clinical Chemistry 1: 329-344, 1955;
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Clinical Chemistry, Vol 1, 329-344, Copyright © 1955 by the American Association for Clinical Chemistry

The Correspondence with Beer's Law for the Optical Density of Stained Protein Patterns on Filter Paper as a Function of Surface Protein Concentration

V. H. Rees 1 and D. J. R. Laurence 1

1 Department of Chemical Pathology, Postgraduate Medical School of London, Ducane Road, London, W. 12, England.

The results obtained here consistently failed to show deviations from Beer's law for optical densities less than 1.4, and the use of scanning as a convenient method for differential protein estimations would appear to be justified. The methods described may enable other workers to make similar tests of the method with a minimum of preliminary development, and some may succeed in obtaining significant failure of Beer's law in their apparatus. The use of apparatus of this type would enable the variation of the deviations with arrangement of the optical system, etc., to be worked out, but the following experimental conditions are already known to lead to significant deviations:

1. Use of dry paper instead of lightly oiled paper.

2. An inadequate light ifiter for the photocell.

3. An illuminated slit too long to be completely covered by the protein pattern.

4. Variations in protein density over the slit, either because the slit is too wide or because the protein pattern has been applied unevenly.

5. Use of low quality ifiter paper containing "pin holes."

6. A dye uptake which is not proportional to protein content of the paper [Martin and Franglen (14)].

The failure of Crook, Harris, and Warren (4) to substantiate Beer's law does not indicate the most general situation for the application of the scanning method.

The application of Beer's logarithmic law to the scanning of dyed protein patterns has been investigated by methods described in detail. No deviations could be found for optical densities less than 1.4 for amidoschwarz 1OB or less than 1.1 for azocarmine B staining. The scanning method can be used for evaluation of protein fractions if care is taken.

Submitted on April 12, 1955




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Copyright © 1955 by the American Association for Clinical Chemistry.