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A convenient, inexpensive organ culture device
ANALYTICAL
BIOCHEMISTRY
A Convenient,
77,
%&.%+l
(1977)
Inexpensive
Organ Culture Device
Organ culture of endocrine glands and of bone is a technique frequently employed to study cellular physiology by radioisotopic methods (l-5). This technique utilizes watch glasses or culture dishes containing suitable inert supports upon which the tissues to be studied are placed. These devices often require substantial space in an incubator if a large number of specimens is studied and require extensive cleaning and sterilization if they are made of reusable glass. We describe a method in which a disposable multichambered plastic device is used, which requires a minimum of space in a incubator and utilizes a very small volume of culture medium. A commercially available brand of plastic dish (Falcon MicroTest II) fitted with a lid contains 96 wells each of which has a volume of 0.4 ml. Before culture, each well is fitted with a sterile cylinder of glass tubing, 6 mm in outer diameter and 8 mm in height. The wells are filled with 0.17-0.18 ml of culture medium. The glass cylinder acts as a support for a circular disk of Millipore filter paper (6.5 mm in diameter; Millipore Co., Type HA) upon which the tissue of interest is placed (Fig. 1). This arrangement allows for the simultaneous exposure of the tissue to the gaseous atmosphere and to the culture medium. When the dishes are covered with the lids and incubated at 37°C in a humidified chamber gassed with air-carbon dioxide (95-5%), evaporation is less
TISSUE
FlklER PAPER
T 1 Rmm
GLASS CYLINDER
-
FIG. 1. Cross-section of well and tissue
support.
540 Copyright All rights
0 1977 by Academic Press. Inc. of reproduction in any form reserved.
ISSN COO32697
SHORT COMMUNICATIONS
541
than 5% after 7 days of culture. This device occupies only 17 in.2 and still allows for culturing up to 96 individual organs or groups of tissues. With the inert glass support, the volume of culture fluid needed is reduced to less than one-half the volume usually required per well. This reduction in the volume of medium used means a decrease in the cost of culture medium, serum, and any radioactive isotopes needed. Furthermore, the reduced volume of medium per well increases the concentration of substances released from the cultured tissues and, in the case of hormoneproducing tissues, allows for measurement of hormones secreted from individual tissues. In addition, it is very easy to plate out multiple experimental media in the various wells and to transfer explants rapidly. Finally, the cost per unit (less than $2.00) is substantially less than that of other glass devices which require a considerable expenditure of labor for their preparation. REFERENCES 1. Strangeways, T. S. P., and Fell, H. B. (1926) Proc. Roy. Sot. London Ser. B 99, 340-366. 2. Wolff, M. E. (ed.) (1961) La Culture Organotypique, Centre National de Recherche Scientifique, Paris. 3. Dawe, C. J. (ed.) (1963) Symposium on Organ Culture: Studies of Development, function and disease, Nut. Cancer Inst. Monogr. 11, l-252. 4. Bingham, P. J., and Raisz, L. G. (1974). Calcif. Tissue Res. 14, 31-48. 5. Messer, H. H., Tuen, S. Y., and Copp, D. H. (1974) Culcif. Tissue Res 16, 89-92.
J. VERA A. LICATA F. C. BARTTER Hypertension-Endocrine Branch National Heart and Lung Institute National Institutes of Health Bethesda, Maryland 20014 Received May 12, 1976; accepted
August
23, 1976
BIOCHEMISTRY
A Convenient,
77,
%&.%+l
(1977)
Inexpensive
Organ Culture Device
Organ culture of endocrine glands and of bone is a technique frequently employed to study cellular physiology by radioisotopic methods (l-5). This technique utilizes watch glasses or culture dishes containing suitable inert supports upon which the tissues to be studied are placed. These devices often require substantial space in an incubator if a large number of specimens is studied and require extensive cleaning and sterilization if they are made of reusable glass. We describe a method in which a disposable multichambered plastic device is used, which requires a minimum of space in a incubator and utilizes a very small volume of culture medium. A commercially available brand of plastic dish (Falcon MicroTest II) fitted with a lid contains 96 wells each of which has a volume of 0.4 ml. Before culture, each well is fitted with a sterile cylinder of glass tubing, 6 mm in outer diameter and 8 mm in height. The wells are filled with 0.17-0.18 ml of culture medium. The glass cylinder acts as a support for a circular disk of Millipore filter paper (6.5 mm in diameter; Millipore Co., Type HA) upon which the tissue of interest is placed (Fig. 1). This arrangement allows for the simultaneous exposure of the tissue to the gaseous atmosphere and to the culture medium. When the dishes are covered with the lids and incubated at 37°C in a humidified chamber gassed with air-carbon dioxide (95-5%), evaporation is less
TISSUE
FlklER PAPER
T 1 Rmm
GLASS CYLINDER
-
FIG. 1. Cross-section of well and tissue
support.
540 Copyright All rights
0 1977 by Academic Press. Inc. of reproduction in any form reserved.
ISSN COO32697
SHORT COMMUNICATIONS
541
than 5% after 7 days of culture. This device occupies only 17 in.2 and still allows for culturing up to 96 individual organs or groups of tissues. With the inert glass support, the volume of culture fluid needed is reduced to less than one-half the volume usually required per well. This reduction in the volume of medium used means a decrease in the cost of culture medium, serum, and any radioactive isotopes needed. Furthermore, the reduced volume of medium per well increases the concentration of substances released from the cultured tissues and, in the case of hormoneproducing tissues, allows for measurement of hormones secreted from individual tissues. In addition, it is very easy to plate out multiple experimental media in the various wells and to transfer explants rapidly. Finally, the cost per unit (less than $2.00) is substantially less than that of other glass devices which require a considerable expenditure of labor for their preparation. REFERENCES 1. Strangeways, T. S. P., and Fell, H. B. (1926) Proc. Roy. Sot. London Ser. B 99, 340-366. 2. Wolff, M. E. (ed.) (1961) La Culture Organotypique, Centre National de Recherche Scientifique, Paris. 3. Dawe, C. J. (ed.) (1963) Symposium on Organ Culture: Studies of Development, function and disease, Nut. Cancer Inst. Monogr. 11, l-252. 4. Bingham, P. J., and Raisz, L. G. (1974). Calcif. Tissue Res. 14, 31-48. 5. Messer, H. H., Tuen, S. Y., and Copp, D. H. (1974) Culcif. Tissue Res 16, 89-92.
J. VERA A. LICATA F. C. BARTTER Hypertension-Endocrine Branch National Heart and Lung Institute National Institutes of Health Bethesda, Maryland 20014 Received May 12, 1976; accepted
August
23, 1976