Immunofluorescent Antibody Technique Research Articles

AN IMMUNOFLUORESCENT STUDY OF SPERM ISOIMMUNIZATION IN INFERTILE WOMEN

SummarySerum from 32 infertile women and from eight control patients was investigated for antisperm activity using an indirect immunofluorescent antibody technique. The results confirmed previous reports of the surface localization of antigens on human spermatozoa. Specific antibody staining was observed on the sperm acrosome and tail; neck and equatorial segment staining was common and non‐specific. Antibodies in the IgM and IgG classes localized on the acrosome, whereas tail staining was exclusively due to IgG antibodies. In seven out of eight sera in which a complement‐dependent sperm immobilization test was positive, an IgG antibody was demonstrated which localized on the sperm tail. There was a poor correlation between positive sperm micro‐agglutination and specific immuno‐fluorescent reactions.

Requirement of Arginine for the Replication of Marek's Disease Herpes Virus

SUMMARY The requirement of arginine for the replication of Marek’s disease herpes virus (MDHV) was studied in Japanese quail embryo fibroblast cell (QEFC) cultures. Arginine was essential for the replication of MDHV in QEFC cultures. The plaqueforming activity of MDHV was inhibited by omission of arginine from the medium but resumed upon addition of arginine in arginine-deprived infections. The MDHV genome survived in a recoverable form in infected cultures kept in arginine-deprived medium for up to 10 days. Arginine was required around or after 12 h following inoculation. Omission of arginine from the culture medium did not affect virus adsorption, penetration, or the synthesis of virus DNA but did prevent the formation of virus structural protein which was detectable using immunofluorescent antibody techniques.

Antigenic deletion in cervical neoplasia

Using an indirect immunofluorescence antibody technique, specific antigen charges have been followed in pre-invasive and invasive cervical carcinoma. Two specific antigenic components of squamous epithelium have been studied, an intercellular surface antigen and a basement membrane antigen. In histologically normal epithelium, intercellular and basement membrane antigens can be shown. Inflammatory conditions, age, pregnancy and cyclic activity do not affect antigenic status. Both intercellular and basement membrane antigens are deleted in frank invasive squamous cell carcinoma of the cervix, but in pre-invasive lesions antigen deletion occurs to a varying extent and is proportional to the increasing severity of the pre-invasive lesions. Using an indirect immunofluorescence antibody technique, specific antigen charges have been followed in pre-invasive and invasive cervical carcinoma. Two specific antigenic components of squamous epithelium have been studied, an intercellular surface antigen and a basement membrane antigen. In histologically normal epithelium, intercellular and basement membrane antigens can be shown. Inflammatory conditions, age, pregnancy and cyclic activity do not affect antigenic status. Both intercellular and basement membrane antigens are deleted in frank invasive squamous cell carcinoma of the cervix, but in pre-invasive lesions antigen deletion occurs to a varying extent and is proportional to the increasing severity of the pre-invasive lesions.

Attempted Application of Avian Viral Antibody Test for Detection of Other Microbial Infections

SUMMARY An indirect immunofluorescence antibody technique was used to test 914 egg yolk extracts from 92 chicken flocks in 12 states against 11 microbial antigens. The almost universal reactivity of the egg yolk extracts with the more complex microbial antigens seems to preclude a practical application of this reaction for the detection of other than viral antibody in avian flocks.

Antibodies to herpes-type virus in nasopharyngeal carcinoma and control groups.

Sera collected from patients with nasopharyngeal carcinoma (NPC), in Taiwan, were titrated for antibodies to herpes-type virus (HTV) in a Burkitt's lymphoma cell line (P3HR-1) by the indirect immunofluorescence antibody technique. The results were compared with those of neighborhood controls matched by age and sex, with patient families and with neighborhood control families. Antibody liters were higher in the NPC cases than in any of the 3 control groups. Dissociations in the frequency distributions of antibody titers in the NPC and healthy control groups were found to be maximum when the limiting value was set at 1:640. There were no significant differences in the frequency distribution among the 3 control groups. Of 117 NPC patients, 40% had anti-HTV titers equal to or greater than 1:640, while such values occurred in only 5, 5, and 6%, respectively, of the neighborhood controls, patient families, and control families. Though not statistically significant, there was difference in the percentages of “sero-positive” cases for the NPC patients by sex—42% for males and 35% for females. There was no sex difference, or it was a little higher in females, in the 3 control groups. The geometric mean titers were 1:242, 1:65, 1:69, and 1:61, respectively, for the NPC patients, neighborhood controls, patient families, and neighborhood control families. There was a difference in the geometric means by sex in the patients, i.e., 1:253 and 1:210, respectively, for males and females, while no differences by sex were noted in the 3 control groups. Ridit analysis and chi square test showed definitely different results between the NPC patients and the 3 control groups. The relative risks indicated that persons with antibody titer of 1:2560 had more than 200 times the NPC risk of those with antibody titer of less than 1:40. There were no indications of household aggregation of high anti-HTV titer (≥ 1:640) in the patients and control families. The significance and implications of these findings are discussed.

A Rapid Diagnostic Aid for Herpesvirus hominis Infections in Dermatology

An indirect immunofluorescent antibody technique is described which can be used to identify rapidly, specifically and accurately herpesvirus hominis infections affecting the skin. The possible applications of this technique as a diagnostic aid of suspected herpetic infections in dermatology are illustrated and discussed. It is suggested that the method should prove useful in the investigation of all vesicular skin lesions in which infection with herpesvirus hominis is suspected. With the advent of anti-viral agents these techniques are likely to become increasingly important.

IMMUNOFLUORESCENT STAINING OF MUCOSAL GLYCOPROTEIN OF HUMAN INTESTINAL MUCOSA

Mucosal glycoprotein was isolated from human intestinal mucosa. It possessed higher sialic acid and sulfate contents than those of the gastric mucosal glycoprotein. The former was, therefore, stainable with Alcian Blue while the latter was not. These two kinds of glycoproteins gave a specific precipitate line against the corresponding antiserum, respectively, on the Ouchterlony assay. Using the direct immunofluorescent antibody technique, the intestinal glycoprotein was demonstrated to distribute in the alcianophilic goblet cells throughout the intestinal mucosa but only faintly demonstrable in the surface epithelium of gastric mucosa.

Immunofluorescent assay of herpesvirus type 1 and type 2 antibodies in rabbit and human sera

The immunofluorescent antibody technique was used to assay type-specific herpes simplex virus antibodies. Antibody responses were studied in three groups of patients: children with primary herpetic infections, neonates with generalized type 2 infection and adults with genital herpes infections. The results were compared with antibody responses seen in rabbits after experimental type 1 and 2 infections. In both rabbits and children with primary herpetic infections there was considerable cross-reaction, while in patients with genital herpes infections and in neonates there was very little cross-reaction.

THE NUCLEAR STAINING PATTERNS OF ANTINUCLEAR FACTORS OBSERVED WITH IMMUNOENZYME ANTIBODY TECHNIQUE

By means of the indirect immunoenzyme antibody technique for the detection of antinuclear factors, the sere from 18 patients with SLE were examined, and nuclear staining patterns observed were classified as follows; diffuse, perinuclear, flecked, fine granular, and thready types. These five types had no relationship to the types of nuclear staining by the indirect immunofluorescent antibody technique. There were no changes in staining patterns under various fixative conditions, and serial dilution of sera did not result in a type different from the undiluted one. Good stainings were obtained when using glutaraldehyde as a bifunctional reagent. As to the sensitivity of the nuclear staining, immunofluorescent antibody technique were seemed to be superior to immunoenzyme antibody technique. The origins of these newly defined types were regarded as due to the factors other than specific nuclear antigens.

56.On the localization of βI_c/IA-and βIE-gulobulin in the liver of chronic active hepatitis by an immunofluorescent antibody technique

56.On the localization of βI_c/IA-and βIE-gulobulin in the liver of chronic active hepatitis by an immunofluorescent antibody technique

Antinuclear Factors, the Antihumanglobulin Consumption Test, and Wassermann Reaction in Psoriatic Arthritis

SummarySera from 50 patients with psoriasis and arthritis, 46 without rheumatoid factor, were studied for the presence of antinuclear factors as determined by the immuno-fluorescentantibody technique (ANF) or the antihumanglobulin consumption test (AGKT). Three out of 48 sera had a positive ANF-reaction, and one out of 44 sera showed a strongly positive AGKT-reaction (+ +). Ten out of 44 sera showed weak AGKT-reactions (+), which are found in sera of normal individuals. No biologically false positive reactions for syphilis were demonstrated among the fifty patients. Thus, the present study showed no special immunological patterns of reaction in psoriatic arthritis.

Production of kidney tissue cultures from African green monkeys, experimentally infected with the causative agent of Frankfurt-Marburg-syndrome.

Green monkeys (Cercopithecus aethiops), experimentally infected with the causative agent of the Frankfurt-Marburg-Syndrome (FMS-virus), were killed after the appearance of disease symptoms, and. nephrectomized. Tissue cultures were set up from the trypsinized kidney tissue. Examination of live tissue culture under low magnification revealed no difference in rate of growth or appearance from the cultures of kidney cells from non-infected monkeys. Intracytoplasmic eosino-philic inclusion bodies could be found invariably in stained preparations of the cells of kidney tissue cultures deriving from infected monkeys, virus particles could be seen by electron microscopy, and viral antigen could be demonstrated in the cells with immunofluorescent antibody technique. That the tissue culture fluids contained infectious virus could be demonstrated by inoculating monkey kidney cell tissue cultures in stationary growth phase and observing the cyto-pathic effect.

Intranuclear crystal formation of poliovirus: Electron microscopic observations

Poliovirus-infected FL cells, incubated at 28°C, were examined by electron microscopy. The immunofluorescent antibody technique was employed for detection of the intracellular viral antigen. Electron micrographs of cells from infected cultures showed both extensive cytoplasmic and nuclear alterations. Virus crystals were observed not only in the cytoplasm but also in the nucleus of cells 14–20 hours after infection. Intracytoplasmic crystals consisted of dense particles, approximately 27 mμ in diameter, suggesting mature virus particles, while intranuclear crystals were composed of empty particles with a less dense central area. Nuclei were stained specifically by fluorescent antibodies as small brilliant foci, suggesting that the intranuclear crystals possess the same antigenicity as the viral antigen. As judged by their size, structure and antigenicity, the intranuclear virus-like particles probably represent an incomplete form of the virus.

Application of immunofluorescent antibody technique in rapid diagnosis of respiratory syncytial virus infection.

Seventy-eight unselected children under the age of 2 years suffering from acute respiratory infections were investigated by the fluorescent antibody technique and a comparison was made with conventional isolation and serological methods. Sixty-nine per cent. of children with bronchiolitis were diagnosed as suffering from respiratory syncytial virus infection on the day of admission by examination of nasopharyngeal secretions. There were 44 children with respiratory syncytial virus infection diagnosed by conventional methods over a month, but by using fluorescent antibody techniques on tissue culture 53% were diagnosed by the second day, 71% by the fourth day, and 82% by the seventh day. The method of choice for a rapid diagnosis of respiratory syncytial virus infection is by examining nasopharyngeal secretions, when 90% of those with this infection can be diagnosed on the day of admission to hospital.

Muscle proteins in the chick myotome examined by the immunofluorescent method

ABSTRACT This work was undertaken to examine by the immunofluorescent antibody technique at which stage of development of the chick embryo the first muscle proteins are formed. In addition the spatial interrelationship of presumptive myoblasts, myoblasts and myotubes was studied. Chick embryos ranging in age from 52 h to 5 days were fixed in Carnoy and transverse sections cut at 3 – 4 μ .The indirect fluorescent method (sandwich technique) was applied using fluor-scent goat gamma-globulin and myosin antiserum. When the specificity of the myosin antiserum was tested by the immuno-electrophoretic technique, antibodies against myosin as well as against actin, adenylic acid deaminase and some other fractions were detected. Consequently, the fluorescent reactions found in the embryo do not specifically indicate the prescence of myosin, but rather that of muscle proteins. The first fluorescent reaction over the myotome was found at stages 17–18, and was restricted to a few cells on the lateral side of the central myotome region. From here the reaction spread rapidly to other cells on the lateral side in a ventral and dorsal direction. Simultaneously with the increase in thickness of the myotome, the fluorescent reaction spread in a medial direction. The cells of the myotome, particularly those in the most dorsal portion, did not begin to show fluorescense until the adjacent dermatome cells had lost their epithelial structure. At stages 24 – 26 the lateral side of the myotome is formed by myotubes with fluorescent myofibrils. In a medial direction are found subsequently heavily fluorescent spindle cells, either in bundles or as individual cells, which in turn are followed by irregular cells either slightly or not at all fluorescent. The latter cells sometimes showed DNA synthesis. Hence, the differentiation from presumptive myoblast to myoblast and from myoblast to myotube proceeds from lateral to medial.

Rapid diagnosis of respiratory syncytial virus infection by immunofluorescent antibody techniques.

Rapid diagnosis of respiratory syncytial virus infection by immunofluorescent antibody techniques.

Autoimmunity in Endocrine Disease

In general there is a parrallel between serum sperm agglutinin titers and the inability of the spermatozoa to penetrate the cervical muscosa. The serum of 67 (3.3%) of 2015 infertile males had titers of 1:32 or higher. Of 416 fertile men and 124 infertile women none had serum with titers of 1:32 or higher. Of the 67 infertile men about 1/3 had semen with normal sperm density and usually autoagglutination; another third had oligospermia with sperm of subnormal motility while autoagglutination was not easily observed; and the remaining third had azoospermia usually due to obstruction of efferent ducts. Of the last group testis biopsies showed normal histology in 15 men and partial abnormality in 2 indicating that sperm autoantibodies did not interfere with spermatogenesis. No cause for antibody formation was observed in more than half of nonazoospermic patients. There was a positive correlation between agglutination and staining of ejaculated spermatozoa by an indirect immunofluorescent antibody technique. Some sera stained both testicular and ejaculated spermatozoa and some only one or the other. Five of 11 men with congenital absence of both the vasa deferentia and seminal vesicles had sperm agglutinins at titers of 1:8 to 1:256 higher levels of gamma A (P=.004) and higher titers of complement-fixing anti-measles virus antibodies (P=.008) than those without sperm agglutinins. Among 25 men whose vasa had been ligated 2 to 20 years previously 17 showed no sperm agglutinins perhaps due to: 1) a variable mechanism of phagocytosis and/or defective digestion of spermatozoa by cells lining the sperm pathway 2) resorption of sperm antigens causing immune tolerance 3) initiation of antibody formation requiring an adjuvant inflammatory effect 4) other antibodies. Corticosteroid and ACTH therapy has not altered the serum titers or autoagglutination of infertile men.

On the incidence of antinuclear factors in rheumatoid arthritis. A comparative study, with special reference to the reaction with polymorphonuclear granulocytes.

SummarySera from 40 patients with definite rheumatoid arthritis were investigated for the presence of antinuclear factors as determined by the immunofluorescent antibody technique, employing different human nuclei as antigen.While altogether 34 of the sera reacted with one or more nuclei 88.2% of the positive sera reacted with the nuclei of granulocytes and only 35% with other human nuclei. One third of the sera reacted selectively with nuclei of granulocytes.The finding that 75% of the sera from patients with rheumatoid arthritis contained ANF reacting with granulocytes was obtained only when granulocytes damaged by freezing and thawing were employed. When fresh, unaltered leukocytes were used, only 30% of the sera reacted.The results indicate that the nuclear reactivity of sera in rheumatoid arthritis is mainly directed against the nuclei of the polymorphonuclear granulocytes. The limited nuclear reactivity of these sera may explain the wide variation of the incidence of ANF previously found in rheumato...

Immunofluorescent antibody technique in the diagnosis of gonorrhoea by direct smears from the female.

Immunofluorescent antibody technique in the diagnosis of gonorrhoea by direct smears from the female.

Coxsackie B viral myocarditis and valvulitis identified in routine autopsy specimens by immunofluorescent techniques

From a survey of 55 routinely autopsied hearts, studied by means of the immunofluorescent antibody techniques, Coxsackie B group virus antigens were found in the myocardium in 17 cases (30.90 per cent) and in both the myocardium and mitral valve in 3 cases (5.45 per cent). A chronic focal, interstitial myocarditis was noted in all 17 cases upon routine histologic study. A high percentage of positive viral antigen was found in hearts of children (75 per cent) and infants (35.71 per cent). The criteria for positive immunofluorescent identification of antigen consisted of intracytoplasmic localization of the fluorescence in the affected myofibers and in fibrocytes of the mitral valves. Chronic Coxsackie virus valvulitis is shown to be present in man in certain types of unexplained chronic valvular heart disease. It is postulated that some instances of chronic valvular disease previously thought to be post rheumatic in origin may represent chronic viral valvulitis. These studies also suggest a possible role of the Coxsackie virus as a cause of some congenital cardiac defects and stillbirths.