An improved photographic system for polyacrylamide gels

ANALYTICAL

BIOCHEMISTRY

An Improved

44, 540-542 (1971)

Photographic DENIS

Department

OLIVER

of Biochemistry,

System AND

for Polyacrylamide

ROGER

University

Received April

Gels

CHALKLEY

of Iowa, Iowa City, Iowa 6,%40 2, 1971

A vexing problem associated with polyacrylamide gel analysis is poor photographic reproducibility for publication. There are several reasons for failure to produce photographic images which are equivalent to those visualized: (1) in order to obtain the high contrast necessary for good reproduction in journals, minor (but important) bands may totally disappear; (2) imperfections in a gel may mar the appearance of bands ; and (S) bands may be closely spaced and exhibit a trace of curvature. While this presents no problem to the eye, which simply compares appropriate, limited parts of the strained regions, the photograph is of course unable to select the most clearly defined regions of a band and the subsequent print may convey little of the evidence adduced by eye. There was a need therefore for a device that could select a small part of each band, that could magnify and intensify this region so that all bands could be clearly visualized, and that would prove amenable to the means of photography most commonly available in the laboratory. This paper describes such a device and documents the photographic improvement which results from its use. THE

APPARATUS

The apparatus is exceedingly simple. The polyacrylamide gels are placed in glass tubes (i.d. 0.6 cm) in a clear solution of acetic acid (0.9 N). The tubes are preferably much longer than the gels. The loaded tubes are then mounted horizontally over a diffuse light source in the apparatus shown in Fig. l-as many as 8 tubes can be conveniently accommodated in this system. An identical tube containing only water1 is then mounted directly over the sample tube beneath the camera lens (75 mm). The camera was an MF-3 fitted with 35 mm film back (Dyonics 35%) * The film used should be of the high contrast variety (Panatomic-X, Kodak: ASA = 30). An improvement in clarity may be noted if an ‘Water

is preferable

to air, ethanol, ethylene glycol, or glycerol. 540

POLYACRYLAMIDE

GEL

541

PHOTOGRAPHY

FIG. 1. Photograph of apparatus with tubes in place. The band straightening apparatus consists of two Lucite blocks (a), 23.5 X 5 X 2 cm with slits 17 mm in width and 39 mm in depth, into which two glass tubes are placed (one over the other) spanning the two blocks. The tube containing the electrophoresis gel (specimen tube-b) is positioned below a tube of water (focusing tube-c). The apparatus is set above a diffuse light source (e.g., an x-ray viewer, d) with the camera positioned directly above. In the figure, the light box has been covered with black paper to provide contrast for photography of the apparatus. The apparatus is set over a diffuse light source (e.g., an x-ray viewer, d) with the camera positioned directly above.

orange filter is used. If a figure is made directly from the first negat’ive, #3 or #4 contrast print paper is preferable. However, if a figure is constructed from a series of photographs and this composition is rephotographed, then optimum resolution will demand that the final picture be on #5 or #6 contrast printing paper. TYPICAL

RESULTS

OBTAINED

WITH

THIS

SYSTEM

Interposition of a tube containing water between sample tube and camera solves the problems discussed in the introduction. Because of refraction of transmitted light by the water tube, the portion of the stained gel observed is confined essentially to the line of contact between the two tubes. Since only that portion of each band nearest the camera is thus photographed, curved bands now appear straight. As a cylindrical lens the water tube introduces no distortion parallel to its axis; the distances between bands are thus not altered in the photograph. Figure 2 shows photographs of typical results we have obtained following polyacrylamide gel electrophoresis of histones from various sources. For comparison, we show t’he appearance of the bands photographed either with or without utilizing the system described above.

542

OLIVER

AND

CHALKLEY

FIG. 2. Effect of band straightening on various band patterns: (a) poor gel; (b) distorted gel; and (c) good gel. In each case the gel photographed using the apparatus described is shown to the right of the control gel. Electrophoresis followed the method described by Panyim and Chalkley (1).

Longer gels are becoming increasingly popular and the occurrence of truly flat bands in these systems is not common. Figure 2a shows a product from a long gel (15 cm). In Fig. 2b we see that even a grossly deformed band pattern can be converted into a form useful for comparison of band mobilities and which at the same time assumes a more esthetically desirable appearance. Finally, it is seen that, if the original gel (Fig. 2c) is of good quality, with flat well-resolved bands, then the improvement obtained with this approach is minimal, though the increase in band intensity may lead to greater contrast in the final photograph. ACKNOWLEDGMENTS We wish to thank our colleagues in this laboratory for their youthful skepticism. This work was supported by grant CA-19371 from the National Cancer Institute of the U. S. Public Health Service, by American Cancer Society (Iowa Division) grant 491, and by an NIH Predoctoral Developmental Biology Training grant 5T91-HO-00152 to one of us (D.O.).

1. PANYIM,

S., AND CHALKLEY,

REFERENCE R., Biochemistry

8, 3972 (1969).