Indirect Immunofluorescence Procedure Research Articles

The ultimobranchial gland of the dogfish Scyliorhinus canicula L. Immunofluorescence cross-reactivities with anti-sera to different calcitonins.

Calcitonin in the ultimobranchial gland of the dogfishScyliorhinus canicula cross-reacts with anti-dogfish, anti-salmon and anti-porcine calcitonin, but not with anti-human calcitonin, using indirect immunofluorescence procedures. These results do not reflect completely expectations derived from the primary structures of the hormones, where known, and from their physiological interrelationships.

Advances in methodology for immunodiagnosis of parasitic diseases

During recent years, significant progress has been made in the development and improvement of procedures for the immunodiagnosis of parasitic diseases. The purpose of this review is to briefly discuss the inherent advantages and limitations of various immunodiagnostic methods currently in general use, to delineate areas in which further research is needed, and to call attention to new technics that may prove to be valuable additions to the present armamentarium of procedures for the immunodiagnosis of parasitic infections. It is well established that the specificity and sensitivity of an immunodiagnostic procedure depend on the quality of the antigen employed. The introduction of better methods for isolating the serologically active antigen components from crude parasite extracts has significantly improved the pathognomonic value of many immunodiagnostic tests. Since the chemical nature of the major specific antigen(s) differs among the various parasites, no single method is universally suitable for antigen fraetionation. Studies illustrating the variety of physicochemical fractionation procedures that have been used successfully for the isolation of specific parasite antigens are noted. In addition, attention is called to the potential advantages of employing exoantigens (i.e., excretions and secretions of living parasites) in immunodiagnostic tests. Procedures most frequently employed for immunodiagnosis of parasitic diseases are discussed in some detail and include: complement fixation; indirect hemagglutination; indirect immunofluorescence procedures using whole organisms as antigens; flocculation tests employing lecithin-cholesterol crystals, bentonite or latex particles as the antigen matrix; card tests using antigen-sensitized lecithin-cholesterol crystals with added charcoal; agglutination reactions; precipitin tests; and hypersensitivity reactions. The advantages and limitations of these procedures are noted and areas in which their use could be extended are suggested. Certain less commonly employed procedures that have shown considerable promise for serodiagnosis or for basic studies on the immunology of parasitic diseases also are discussed. These include immunoelectrophoresis, the recently developed soluble antigen fluorescent antibody procedure, in vitro histamine release by in vivo-sensitized leukocytes, passive cutaneous anaphylaxis, conglutinating complement absorption procedures, and serological adhesion (immune adherence) reactions. Problems associated with the wide variations of methodology employed in various diagnostic laboratories are noted and the need for improved standardization of antigens and technics is emphasized.

Genetic analysis of simian virus 40: II. Comparison of seven dilution end-point titration microassays

Mammalian cells have been successfully cultured in volumes of 0.002 to 0.012 ml. Using these microcultures and an indirect immunofluorescent staining procedure, we have developed a dilution end-point titration micromethod for use with simian virus 40. This titration procedure is faster, as accurate, and less susceptible to contaminating infection than the presently available plaque assay method. The primary advantage of the technique, however, is that it should allow an investigator to carry out extensive physiologic and genetic analyses of viruses and cells on a modest budget and without technical assistance.The kinetics of SV40 tumor antigen and virion antigen synthesis at various temperatures, and the propagation of single virions were studied in microcultures and found to be similar to those determined by conventional methods.A replica-plating technique for virus has been developed that allows one person to screen more than 1000 single virus clones per week.

Genetic analysis of simian virus 40: I. Description of microtitration and replica-plating techniques for virus

Mammalian cells have been successfully cultured in volumes of 0.002 to 0.012 ml. Using these microcultures and an indirect immunofluorescent staining procedure, we have developed a dilution end-point titration micromethod for use with simian virus 40. This titration procedure is faster, as accurate, and less susceptible to contaminating infection than the presently available plaque assay method. The primary advantage of the technique, however, is that it should allow an investigator to carry out extensive physiologic and genetic analyses of viruses and cells on a modest budget and without technical assistance. The kinetics of SV40 tumor antigen and virion antigen synthesis at various temperatures, and the propagation of single virions were studied in microcultures and found to be similar to those determined by conventional methods. A replica-plating technique for virus has been developed that allows one person to screen more than 1000 single virus clones per week.

Substitution of egg yolk for serum in indirect fluorescence assay for Rous sarcoma virus antibody.

ConclusionsSubstitution of egg yolk for the mother hen's serum does away with the cumbersome procedure of bleeding the birds for antibody tests. The level of RSVA in yolk reflects the serum antibody level but is usually somewhat lower. Furthermore, substitution of egg yolk for the mother hen's serum in an indirect immunofluorescence procedure provides a simple, rapid, reliable and relatively inexpensive testing system. Flocks of birds can be easily examined and in most cases the test results are available the same day.

Soluble antigens for immunofluorescence detection of Histoplasma capsulatum antibodies.

Soluble antigens from Histoplasma capsulatum in the mycelial and yeast phase were purified by gel filtration, fixed onto paper discs, and employed in an indirect immunofluorescence procedure to detect antibody in sera from individuals infected with H. capsulatum. The elution patterns of crude histoplasmin passed through Sephadex G-200 revealed two minor peaks of protein showing immunofluorescence, complement fixing, and precipitating-antigen activity. A large peak containing the pigment and other low molecular weight materials showed no serological activity. A polysaccharide antigen obtained from fragmented, deproteinized yeast-phase cells was reactive in the fluorescent-antibody test but showed no antigen activity in complement fixation or precipitin tests. Although certain sera from culturally proven cases of blastomycosis, coccidioidomycosis, and cryptococcosis reacted with the purified Histoplasma antigens, preliminary evaluation indicated that the immunofluorescence technique may be of value as a screening procedure for the serodiagnosis of histoplasmosis.

Soluble Antigens for Immunofluorescence Detection of Histoplasma capsulatum Antibodies

Soluble antigens from Histoplasma capsulatum in the mycelial and yeast phase were purified by gel filtration, fixed onto paper discs, and employed in an indirect immunofluorescence procedure to detect antibody in sera from individuals infected with H. capsulatum. The elution patterns of crude histoplasmin passed through Sephadex G-200 revealed two minor peaks of protein showing immunofluorescence, complement fixing, and precipitating-antigen activity. A large peak containing the pigment and other low molecular weight materials showed no serological activity. A polysaccharide antigen obtained from fragmented, deproteinized yeast-phase cells was reactive in the fluorescent-antibody test but showed no antigen activity in complement fixation or precipitin tests. Although certain sera from culturally proven cases of blastomycosis, coccidioidomycosis, and cryptococcosis reacted with the purified Histoplasma antigens, preliminary evaluation indicated that the immunofluorescence technique may be of value as a screening procedure for the serodiagnosis of histoplasmosis.