Immunofluorescence Labeling of Calcium Pump

adult skeletal muscle and embryonic heart

Immunofluorescence was used to lable the chicken calcium ATPase. This is a sinlge adult slow-twich skeletal muscle fiber grown in tissue culture. You can see that the same caclium ATPase has a very different subcellular distribution in a muscle compare to a skin cell.

Figure 1. Adult slow-twich skeletal muscle grown in cluture. Skeltal muscles are multinucleated and eight nuclei in a row that are labled in this photomicrograph. The more abundant lableing is striated, or striped, because this calcium ATPase is located in the muscle cell's ER, which in muscle cells is called the sarcoplasmic reticulum (SR). The SR striations are in phase with the striated thick and thin myosin and actin proteins that form the sarcomeres. Two other skeletal muscle cells are visible below this one that is in focus. Photograph used with permission from Dr. Doug Fambrough, Johns Hopkins University. (See Kaprielian et al. 1996. Journal of Cell Science 109: 2529-2537.

In this example, the same antibody was used to label calcium ATPase in developing chicken embry hearts. The first diagram shows what the heart looks like at this stage of development. These diagrams are classic embryology figures dating back over 50 years.

Figure 2. Two line drawings showing the position and shape of the developing chicken embryonic heart. The embryos are at developmental stages 11 (panel A) and early13 (panel B) which are separted by only a couple hours. Adapted from Patten and Kramer (1949).

The two photomicrographs below show brightfield illumination (left) and fluorescence illumination of the same embryonic heart. As you can see in this color photomicrograph, the calcium ATPase is already expressed in cardiac cells.

Figure 3. Photomicrographs of an embryonic chicken heart in bright field (left) and immunofluorescence (right) illumination. Since color film was used, the secondary antibody appears green instead of white in the photograph. Photographs taken by Jeffery Cromartie.


List of Methods

© Copyright 2001 Department of Biology, Davidson College, Davidson, NC 28036
Send comments, questions, and suggestions to: