<b>ENZYME IMMUNOASSAY SYSTEM FOR MEASUREMENT OF THREE FORMS (MM, MB AND BB) OF RAT CREATINE </b><b>KINASE </b>

Abstract

A highly sensitive sandwich-type enzyme immunoassay system for the assay of rat creatine kinase (CK) MM, MB and BB isozymes was developed by use of [5'-D-galactosidase from Escherichia coir' as label and the purified antibodies raised in rabbits by injecting CK-MM or CK-BB. The antibodies specific to each subunit were purified from the immunoglobulin G (lgG) fractions of antisera by immunoaffinity chromatography with a column of respective antigen (CK-MM or CK-BB)-coupled Sepharose. The CK-MM and CK-BB assays were performed with the respective antibody F(ab')2 fragments-immobilized polystyrene balls and the same antibody Fab' fragments labeled with ,6'-D-galactosidase from Escherichia c0h'. The CK-MB was assayed with the polystyrene balls with either antibody (anti-CK-M or anti-CK-B) and another antibody (anti-CK-B and anti-CK-M, respectively) labeled with galactosidase. The minimum detection limit of each assay was 1 to 10 pg or (l0 to 100 amol) per tube. The assays for CK-MM and CK-BB were cross-reacted about l2°/0 with CK-MB, but the assay for CK-MB was specific showing no cross-reactivity with and no interference by CK-MM and CK-BB under the limited conditions. Therefore, the concentrations of CK-MM and CK-BB were able to estimate by subtracting the cross-reacting CK-MB in the samples. The concentrations of three forms of CK in various rat tissues were determined by the present assay system. High concentrations of CK-MM were observed in the heart and tissues composed of striated muscle or chondrocytes. CK-MB was also present at relatively high levels in the above-mentioned tissues, while CK-BB was mostly located in the central nervous system and tissues composed of smooth muscle. These results are in line with the previous reports on the distribution of CK isozymes in human tissues.

Nordihydroguaiaretic acid (NDGA) blocks the differentiation of C2C12 myoblast cells

“…Concentrations of CKM, a representative differentiation marker in muscle cells, in extract of cells were determined with a specific immunoassay employing rat CKM purified from skeletal muscle as the standard (Kato and Shimizu, 1987).…”

Localization of brain type creatine kinase in kidney epithelial cell subpopulations in rat

“…A means of attacking this problem is to study homologous systems to distinguish adaptive features from those related to a common history. Several studies have shown that the identification of putative homologues in hervous systems is feasible 2 -4 although dissimilar characteristics, in the absence of developmental evidence, could obscure the homologous relationship of neurones 5,6 There is now considerable evidence to support the conclusion that the insect wing is monophyletic and that it originated as a primitively articulated wing appendage 7. Although the evolutionary precursor to the behaviour of flight is still unclear 8 -1i, similarities in the structure and organization…”

Immunohistochemical study on the development of extraocular muscles. I. Rat.

“…Horizontal and sagittal sections of 5,om thickness of the head were prepared and were immunostained after avidin-biotin-peroxidase complex (ABC) method of Hsu (9) using CK-B, CK-M, j3-enolase and NSE antibodies. The preparation procedures of the antibodies used in the present study are described in other papers (11,13,14). For controls, sections were incubated with normal rabbit serum.…”