Springer Online Journal Archives 1860-2000
Summary A highly sensitive sandwich ELISA for parvalbumin (PA), based on a fluorometric detection system, was developed. This assay detected PA concentrations as low as 20 pg/ml (2 pg per assay) and was used for measuring PA contents in fragments of single muscle fibers isolated from freeze-dried 100–150 μm thick cross sections. The fibers were typed according to their histo-chemically assessed mATPase in parallel cross sections. Type I fibers from rabbit tibialis anterior (TA) and vastus lateralis (VL) muscles contained extremely low PA concentrations (2–5 μg/g w.wt.). Type IIA fibers displayed slightly higher values with mean values of 17 and 29 μg/g w.wt. (range 5–65) in TA and VL, respectively. Much higher PA concentrations were found in type IIB fibers with wide ranges from 75–1150 μg/g w.wt. in TA and 440–1370 μg/g w.wt. in VL. Whereas the IIB fibers of the TA displayed a continuum, two subgroups were distinguishable according to their PA contents (means of 590 and 1230 μg/g w.wt.) in VL. Possibly, the population with the lower PA content which was histochemically defined as type IIB in the present study, corresponds to fiber type IID. The finding that PA is predominantly present in type IIB fibers was also confirmed by the parallel decay of PA and type IIB fibers during chronic low-frequency stimulation. The use of freeze substitution, or alternatively, of freeze-drying, made it possible to demonstrate PA immunohistochemically without artifacts and to evaluate the staining intensity by microphotometry. Performing measurements on the same fibers with the two methods, it was possible to establish a relationship between immunohistochemical staining intensity and PA concentration. This correlation can be used to assess PA contents by evaluating immunohistochemical staining intensities in comparative measurements within the same section.
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