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A rabbit antibody was used to detect factor VIII-related antigen in tissue sections by means of the indirect immunofluorescence technique. Interaction between antibody and antigen was found on vascular intima associated with endothelial cells in all tissues studied. Since there is a close relationship between factor VIII-related antigen and biological factor VIII activity in normal plasma one may draw conclusions about the distribution of factor VIII in tissues.

Two distinct and separate patterns of epidermal cytoplasmic fluorescence were seen with indirect immunofluorescence on human normal skin cyrostat sections. 1. Upper epidermal cytoplasmic (UEC) . This fluorescence was seen only in the upper and middle epidermis. The epidermal cytoplasm fluoresced brightly except for that of the basal cell layer which was negative and contrasted sharply with the rest of the epidermal cytoplasm. No cytoplasmic fluorescence was seen in the dermis. This pattern was found with sera from patients with bullous and nonbuflous diseases. 2. General epidermal cytoptasmic (GEC) . This pattern differed from UEC in that the entire epidermis fluoresced equally intensely. Furthermore, strong cytoplasmic fluorescence was noted in the dermis. This pattern was seen in live patients with nonbullous diseases and in one patient with bullous pemphigoid. Cytoplasmic fluorescence was seen on human spleen touch imprints with sera producing both these patterns, but no cytoplasmic fluorescence occurred on human huffy coat peripheral blood smears.

Fifteen separate and distinct nuclear immunofluorescent patterns have been identified by indirect immunofluorescence employing human spleen touch imprints as substrate. The patterns may be classified into three groups. 1. Patterns of major diagnostic importance: This group includes the peripheral pattern seen in systemic lupus erythematosus (SLE), the thready pattern seen most commonly in LE, occasionally in scleroderma, but not in rheumatoid arthritis; the speckled pattern mainly confined to scleroderma and Raynaud's disease; and nucleolar fluorescence found almost exclusively only in scleroderma. 2. Patterns of less specificity but still of helpful diagnostic importance: A pattern of large speckle-like threads was seen mainly in LE while a nucleolar-like speckled pattern was generally not seen in LE. 3. Patterns not associated with any specific disease: These patterns have to be differentiated from those specifically associated with certain diseases. Malignancy must be strongly suspected when unexplained persistently positive tests are found.

The utilization of radioimmunoabsorption (RIA) for determination of the contents of specific antibody of fluorescent conjugates is described. Samples of seven fluorescent polyspecific anti-human immunoglobulin conjugates along with two controls were labelled with 125I and subsequently absorbed onto IgG-coupled Sepharose beads. Specific anti-IgG antibody concentrations could be calculated by measurements of the gamma emission from the immunoabsorbent. The values obtained were compatible with results achieved by investigation of conjugate performance in an indirect immunofluorescence system using a standard anti-nuclear factor serum.

Interstitial nephritis in rats immunized with heterologous tubular basement membrane. Interstitial nephritis developed in Brown Norway (BN) and Lewis/BN (L/BN) F 1 hybrid rats within two weeks after immunization with bovine cortical tubular basement membrane (TBM) in complete Freund's adjuvant (CFA) or pertussis adjuvant (PA). The histologic lesion began as a polymorphonuclear leukocyte infiltrate and quickly progressed to a primarily mononuclear cell infiltrate with tubular atrophy and tubular epithelial cell degeneration, interstitial and periglomerular fibrosis, and only minimal glomerular abnormalities. It was most severe in BN rats immunized with bovine TBM in PA. Direct immunofluorescence of kidneys revealed linear deposits of IgG along TBM of all proximal tubules and 10 to 20% of distal tubules, and occasionally along the Bowman's capsule. Indirect immunofluorescence revealed anti-TBM antibody in serum and renal eluates. There were no anti-GBM antibodies. Only minor abnormalities of renal function were detected. None of ten additional rat strains similarly immunized developed nephritis. Two strains, F344 and August, did not produce antibody in response to immunization; three strains, Lewis, Wistar-Furth (W/F) and Maxx, developed antibodies but did not possess an antigen on their TBM cross-reactive with the immunizing bovine TBM; and five strains, ACI, Buffalo, Wistar, W/F (fz) and DA, had only small quantities of anti-TBM antibody deposited in their kidneys and did not develop nephritis. Nephrite interstitielle chez des rats immunises avec de la membrane basale tubulaire heterologue. Une nephrite interstitielle se developpe chez les rats Brown Norway (BN) et les hybrides Lewis/BN (L/BN) F 1 en deux semaines apres l'immunisation avec de la membrane basale tubulaire corticale bovine (TBM) dans de l'adjuvant de Freund complet (CFA) ou de l'adjuvant de pertussis (PA). Les lesions histologiques commencent par une infiltration de polynucleaires et deviennent rapidement un infiltrat de cellules mononuclees associe a une atrophie tubulaire avec une degenerescence des cellules epitheliales tubulaires, une fibrose interstitielle et periglomerulaire et des anomalies glomerulaires minimes. Ces images sont plus severes chez les rats BN immunises avec la TBM bovine dans le PA. L'immunofluorescence directe des reins montre des depots lineaires d'IgG le long des TBM de tous les tubes proximaux et de 10 a 20% des tubes distaux et parfois le long de la capsule de Bowman. L'immunofluorescence indirect revele des anticorps anti-TBM dans le serum et les eluats de rein. Il n'y a pas d'anticorps anti-GBM. Seules des anomalies mineures de la fonction renale sont observees. Aucune des dix autres souches de rats immunises n'a developpe de nephrite. Deux souches, F344 et August, n'ont pas produit d'anticorps en reponse a l'immunisation; trois souches, Lewis, Wistar-Furth (W/F) et Maxx, ont produit des anticorps mais n'avaient pas d'antigenes sur leurs TBM aui reagissent avec la TBM bovine immunisante; cinq souches, ACI, Buffalo, Wistar, W/F (fz) et DA, avaient seulement de petites quantites d'anticorps anti TBM deposees sur leurs reins ent n'ont pas developpe de nephrite.

In this study, I used an Olympus single beam microfluorimeter which was modified from the multi-purpose single beam microspectrophotometer according to my specifications.This instrument was used to the sample the accuracy of localization of mouse lens crystallins by direct and indirect immunofluorescence methods. Some of the problems involved in microfluorimetry include: the necessity of the constant intensity of the exciting light, the choice of the filter system and the spot exciting method.From a practical point of view it was concluded that the indirect method of immunofluorescence proivdes a more accurate localization of each lens crystallin than the direct method.

Summary U.v. irradiation of SH-RP cells, a lymphoblastoid cell line carrying Epstein—Barr virus, which was derived from a patient with acute myeloid leukaemia, retarded cell growth but significantly enhanced the percentage of cells containing virus-capsid antigen (VCA) as demonstrated by indirect immunofluorescence. The effects of u.v. irradiation on SH-RP cultures were reversible under normal conditions of medium replenishment every four days. Starving the cultures did not modify these effects.

SummaryThe indirect immunofluorescence test, employing egg yolk extract rather than hen's serum, should offer an important additional tool for the detection of certain microbial antibodies in domestic and wild birds.

The variant 3ct of PARA-adenovirus 7, which was able to synthesize SV40-T antigen in cytoplasm, induced tumors in newborn hamsters. The neoplasms were either undifferentiated adenovirus tumors or pleomorphic sarcomas. Several cell lines were obtained from the tumors: some had the cytomorphology typical of adenovirus or SV40-transformed cells, while others showed ribbon-cell formation. Only the cells derived from the “early” tumors displayed adenovirus T antigen (s); in these cells the SV40 T antigen had a diffuse, granular pattern and was mainly localized on the perinuclear region. In the other cell lines no adenovirus T antigen was shown by the indirect immunofluorescence test; the SV40 T antigen was present in the nuclei, either with the regular pattern of SV40-transformed cells or in the form of faint and sparse fluorescent spots. Le mutant 3cT du PARA-adenovirus 7, qui peut synthetiser l'antigene T du SV40 dans le cytoplasme, a induit des tumeurs chez les hamsters nouveau-nes. Les neoplasmes etaient des tumeurs a adenovirus indifferenciees ou des sarcomes polymorphes. Plusieurs souches cellulaires ont ete obtenues a partir des tumeurs: certaines avaient une cytomorphologie typique des cellules transformees par le SV40 ou les adenovirus, alors que d'autres se caracterisaient par des syncytiums en ruban. Seules les cellules derivees des tumeurs “precoces” contenaient le ou les antigenes T des adenovirus; dans ces cellules, l'antigene T du SV40 est granuleux et diffus, et principalement localise dans la region perinucleaire. Dans les souches, aucun antigene adenoviral T n'a pu ětre decele par immunofluorescence indirecte; l'antigene T du SV40 etait present dans les noyaux, soit sous l'aspect habituel des cellules transformees par le SV40, soit sous forme de fables points lumineux clairsemes.

The use of alpha-fetoprotein for the detection of hepatocellular carcinomas in two adult patients and a newborn infant is presented. The use of alpha-fetoprotein for monitoring the responses of the hepatomas to surgical resection, irradiation, and chemotherapy is described. Alpha-fetoprotein is illustrated in a hepatocellular carcinoma by indirect immunofluorescence.

SummaryThe author has studied 478 sera with the indirect immunofluorescence method and the classical serological methods (Bordet-Wasserman, Reiter, Kahn, V.D.R.L.) and Nelson’s test (TIT). The results of all these agreed for 442 sera (92,4 %), i.e. for 109 posittive and 334 negative sera. But in 36 cases (7,6 %) the results of the immunofluorescence disagree with those of the classical serology or TIT : the IF results were positive 24 times, negative 12 times. But in 33 cases, the IF raction agreed with the clinical diagnosis or with the TIT.In three sera only, the IF reactions seemed to be wrong. They were sera no. 7 (IF negative), no. 27 (IF positive ++) and no. 23 (IF positive + [+]). Taking into account the TIT and eventually the clinical symptoms, the results of the IF test are correct in 475 cases out of 478, and may be false in the 3 other Cases (i.e. 0,3 % of the total).Among the tests of the classical serology, when the IF test is positive, it is the V.D.R.L. test which is most often also positiv...

Antibodies to six trachoma-inclusion conjunctivitis (TRIC) antigens were measured by indirect immunofluorescence in matched specimens of serum and tears from individuals with chronic trachoma. Antibodies (to TRIC antigens type A and/or C) were present in the serum of 78% of Tunisian children, six to eight years old, with severe trachoma and in the tears of 59%. In contrast, antibodies (to TRIC antigens type B, D, and E) were found in the serum and tears of only 6% of Navajo children of similar age with mild trachoma. Among Navajo students 13-20 years old with mild trachoma, 48% had antibodies (to antigens B, D, and E) in serum, but antibodies in tears were very rare. These were usually of lower titer than those in serum, were directed at the same TRIC antigenic types, and paralleled the severity of clinical activity in TRIC infections. The role of lachrymal antibodies in resistance to TRIC infection is not apparent.

Antigens synthesized in LLC-MK 2 cells infected with SA7 were described using indirect immunofluorescence. When cells were reacted with sera from tumor-bearing hamsters, “T” antigens were observed as early as 10 hr post-infection (PI). In certain cells a second type of exclusively intranuclear fluorescence was observed at 16 hr PI. Specific antiviral sera reacted with viral structural antigens approximately 15–16 hr PI. Sera from hamsters immunized with density gradient-purified virus reacted with both “T” antigens and viral antigens.

The endocrine polypeptide (APUD) cells responsible for the production and storage of secretin have been demonstrated, in canine duodenum, by the application of an indirect immunofluorescence technique, using anti-pure porcine secretin, to carbodiimide-fixed cryostat sections. The cells are identified, by the use of parallel cytochemical and ultrastructural preparations, as the small granular S cells. These are essentially restricted to the transitional zone of the duodenal mucosa, with whose lumen their apical projections probably always make contact. There is no need to alter the title of the cells since S can now stand for secretin instead of for small.

Resume— Dans le but d'etudier la localisation cytologique de I'AChE dans divers tissus de Gymnote, nous avons prepare un antiserum anti-AChE en utilisant une preparation commerciale partiellement purifiee, contenant une forme globulaire de l'AChE Gp. Cet antiserum reagit dans le test de double diffusion avec l'enzyme purifieGp, mais egalement avec la forme globulaire spontanee Gb, ainsi qu'avec les trois especes ‘natives’ asymetriques A, C et D. Nous avons mis en evidence l'existence de ces quatres formes Gb, A, C et D dans des extraits de cerveau, de moelle epiniere et de muscle de Gymnote. La forme globulaire Gb est particulierement abondante dans la moelle epiniere et a un moindre degre, dans le cerveau. La distribution des espees est identique dans le muscle et l'organe electrique. Grâce a notre antiserum, nous avom Iocalise par immunofluorescence indirecte l'AChE dans les terminaisons nerveuses sur le muscle, dans le cytoplasme de certains neurones situes dans le centre moteur de la decharge (moelle epiniere) et au voisinage du noyau central de la decharge (cerveau). Abstract— An antiserum, anti-AChE, has been obtained from a partially purified commercial AChE (Gp form) of the electric organ of Electrophorus. This form isverysimilar toa known AChE obtained by tryptic digestion of the ‘native’ forms A, C and D. In a gel double-diffusion test, this serum precipitates the four molecular types of AChE (A, C, D and globular or G form) of the electric organ, which justifies the use of serum anti-Gp for the localization of the native forms A, C and D of the electric organ. Forms analogous to the native forms A, C and D of the electric organ have been found in muscle, in spinal cord and in the brain of the electric eel. However, in the spinal cord and in the brain, a globular (Gp) form is also found in significant amounts. The immunoserum has permitted us to localize by indirect immunofluorescence AChE in muscle motor nerve endings and in the cytoplasm of neurons in nuclei of the spinal cord and brain concerned with the discharge of the electric organ.

Serial blood samples collected from 56 children living in a closed infants' home were examined for antibodies to four human herpesviruses in order to obtain some information on the spread of herpesviruses in an infants' community. Maternal antibodies were detected to all these viruses; however, serology only revealed the presence of the Epstein-Barr virus (EBV) and the cytomegalovirus (CMV) in the course of the observation period. The incidence of antibodies to EBV and CMV increased with the age of the infants and the length of their stay in the infants' home. After 13–14 months of stay, about 90 per cent of the infants possessed antibody against the viral capsid antigen (VCA) of EBV, as demonstrated by the indirect immunofluorescence test, and about 70 per cent possessed CMV antibody, as demonstrated by the complement-fixation test. The antibody against the S antigen of EBV was detected rarely and considerably later after the infection than the VCA antibody.

Cells were separated from tissue specimens of human malignant melanoma and grown in tissue culture on coverslips. After 8 days of growth the explanted cells were fixed and stored until used. Sera from 15 patients with malignant melanoma were reacted with the fixed melanoma cells and a variety of other cells. The indirect immunofluorescence method was used to test for presence of antibodies. Sera from all 15 melanoma patients reacted with fixed melanoma cells giving intense fluorescence of intracytoplasmic granules. The fluorescent material appeared to be independent of melanin granules. Homogenate of fresh melanoma but not of other neoplastic or normal tissue absorbed out the anti-melanoma antibodies. The human antimalignant melanoma antibodies did not react with live malignant melanoma cells or with fixed or live cells from benign nevi or fixed guinea pig kidney, human kidney or HeLa cells. The evidence suggests that these antibodies are unable to penetrate living cell membranes.

An indirect immunofluorescence test using small discs (0.4 cm in diameter) cut from blood smears on filter paper is described. Discs plus three drops of phosphate-buffered saline are laid on antigenic areas on slides, which are then incubated. During incubation, eluted antibodies react with antigen. The method was tested with 10 eluates from healthy persons and with 30 eluates from three groups of 10 patients with Chagas’ disease, schistosomiasis Mansoni and South American blastomycosis, respectively. Sera from the same blood samples were tested also by conventional indirect immunofluorescence tests. Both tests gave similar results. The method proposed is useful as a rapid qualitative screening test when dealing with a large number of samples.

Rabbit anti-mouse brain-associated θ (BAθ) serum has been shown to be cytotoxic to mouse T cells. After suitable absorption this serum can also be used for indirect immunofluorescence. The good correlation between the results of immunofluorescent and cytotoxic tests, and the effect of thymus deprivation, suggests that this serum is a convenient means of specifically visualizing mouse T cells.

The SeroMatic system (automated slide-processing equipment) was evaluated for an indirect immunofluorescence test for syphilis. A study of 6,278 sera examined for the fluorescent treponemal antibody test by manual and automated methods revealed agreement of 98.2% for 2,505 VDRL-positive sera and 99.8% for 3,773 VDRL-negative sera. The reactivity rate of the FTA-ABS was 36.2% and that of AFTA was 35.3% for VDRL-positive sera, whereas with VDRL-negative sera the reactivity rates of FTAABS and AFTA were 12.77% and 12.53%, respectively. Excellent agreement between the two tests indicates that the FTA-ABS test can be automated successfully for screening large numbers of serologic specimens for syphilis.

Summary The normal responses of rubella-specific IgM and IgG antibodies were studied in eight patients with proven rubella by the indirect immunofluorescent technique. To evaluate the detection of rubella-specific IgM for diagnosis, a “blind” study of sera from 13 other patients with proven rubella was carried out and the method proved to be 100 per cent, reliable. Its sensitivity was compared with virus isolation and with direct immunofluorescent staining of virus antigen in infected cells from throat swabs.

Properdin was demonstrated by indirect immunofluorescence in glomeruli of six of seven patients with lupus nephritis, two of whom had membranous lupus nephritis, one focal proliferative lu...

Human thyroid primary, secondary and tertiary monolayer cultures with a low concentration of organ and non-organ specific antigens on the surface of their cells were incubated with different does of phytohaemagglutinin P (PHA). PHA concentrations of 100–500 μg/ml of culture fluid were found to stimulate the appearance of organ-specific and some not specified non-organ-specific antigens that were not demonstrable in unstimulated cultures. The effect was demonstrated with the mixed haemadsorption radial diffusion disc test (MHT) with the mixed haemadsorption isotope test, and with indirect immunofluorescence tests on monolayer cultures grown on cover slips and on suspensions of cells dispersed from monolayer cultures. Whether the stimulatory effect is due to a stimulation of some restricted cell functions or merely to a general stimulation of the cell metabolism that is known to occur, remains to be investigated.

The sequential development of the CELO-specific T and virion antigens was studied in lytically infected chick kidney cells using the indirect immunofluorescence technique. The T antigen was detected 6 hours after infection, and rapidly increased until a peak was reached at 20–24 h. The V antigen could not be detected until 20 h after infection. The V antigen, but not the T antigen, was inhibited by cytosine arabinoside. Both antigens were inhibited by cycloheximide. After storage in liquid nitrogen for 3 weeks, the T antigen in fixed cells could not be detected by indirect immunofluorescence tests. Le Developpement des Antlgfines V et T du Virus Celo Dans les Cellules Renales de Poulet Lytiquement Infectees Le diveloppement siquentiel des antigines V (virioniques) et T speicifiques du virus CELO ete par immunofluorescence indirecte sur les cellules renales de poulets lytiquement infectkes. L'antiment T a eete dicelk 6 heures apres infection et a rapidement prolifere jusqu'a ce qu'un maximum soit atteint entre la vingtisme et favingt-quatrieme heure. l'antigene V n'est apparu que 20 heures apres l'infection. Contrairement a l'antigene T, l'antigene V a ete inhibe par la cytosine arabinoside. Les deux antigenes out ete inhibes par le cycloheximide. Les tests d'immunofluorescence indirecte n'ont pas permis de deceler d'antigene T dans les cellules fixees conservees pendant trois semaines dans de l'azote liquide.

Tumor cells grown as monolayers in microtiter plates are used for the detection of surface antigens by indirect immunofluorescence. A procedure has been developed to keep the cells firmly attached during all reactions and washings. The test is read directly from the plates under a fluorescence microscope. A positive reaction is characterized by a coherent fluoresceinating ring encircling the cell membrane.

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By an indirect immunofluorescence technique antinuclear antibodies were detected transiently, and in low concentrations, in the sera of fourteen of twenty-one patients with infectious mononucleosis. Antinuclear antibodies of both the IgG and IgM variety were encountered. Storage of sera with antinuclear antibodies at 4°C resulted in cryoprecipitation with concomitant loss of antinuclear activity in the supernatant and its appearance in the cryoprecipitates. The antinuclear antibodies encountered did not cause the L.E.-cell phenomenon, nor did they precipitate with a variety of nuclear antigens. The relationship of this finding to idiopathic autoimmune disease requires further study.

An indirect immunofluorescence technique using unfixed colonies is applied to human T-mycoplasmas. The method is found to possess the same degree of specificity as the growth-inhibition test and to be well suited for the identification and classification of T-myco-plasmas. The sensitivity is too low for the detection of antibodies in patient sera. A modification of the indirect haemagglutination test applicable to T-mycoplasmas was less specific, but owing to its sensitivity it can be recommended as a supplement to the metabolic-inhibition test in studies of antibodies in patient sera. The polyacrylamide-gel electrophoresis appeared to be unsuitable for the identification and classification of human T-mycoplasma serotypes.

An outbreak of scrub typhus occurred in Chinese Army personnel in the eastern part of Taiwan during 1970. This is the first outbreak of this disease documented on the main island of Taiwan since 1932. Of 21 hospitalized patients examined during the convalescent stage, 19 had antibody titers of from one to 640 to one to 10,240 as measured by indirect immunofluorescence to the Karp, Kato and Gilliam stains of Rickettsia tsutsugamushi. All were males of from 35 to 51 years and all had high fever by history. Eleven patients had eschars, six with typical black necrotic scabs. In addition, serologic evidence of prior scrub typhus infection was found in 59 of 241 other men from the two military companies involved in the outbreak. All men had been working clearing an area in hills 300 to 500 meters high. This land was originally cleared for agricultural use, but had been abandoned over 4 years previously. Rickettsia tsutsugamushi was recovered from rodents and chiggers (Leptotromibidium deliense) captured in the area.

A high proportion of H2 antigen-activated thymus-derived thoracic duct lymphocytes stained positively with rabbit anti-mouse immunoglobulin reagents as detected by indirect immunofluorescence. In view of the specificity of the reagents used and the fact that T cells from other sources e.g., thymus, were not stained by this technique, it was concluded that the material detected on H-2 antigen-activated thymus-derived thoracic duct lymphocytes was indeed immunoglobulin. When H-2 antigen-activated thymus-derived thoracic duct lymphocytes were cultured in vitro at 37 °C for 18 hr, with or without prior treatment with chymotrypsin, surface immunoglobulin could no longer be detected on the cells. This suggested that immunoglobulin had not been synthesized by the cells but absorbed from elsewhere.

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.

The presence of thyroglobulin in intra- and extrathyroidal lymph vessels of TSH-stimulated guinea pigs was demonstrated immunohistochemically. Guinea pig thyroglobulin was purified by gel filtration on Sephadex G-200 and tested by the double diffusion method and by polyacrylamide gel electrophoresis. Antibodies to guinea pig thyroglobulin were obtained in rabbits by immunisation of the animals with this antigen. Indirect fluorescence staining was performed on formol fixed paraffin embedded tissue with rabbit anti-guinea pig thyroglobulin serum in the first step and fluorescein-labelled antirabbit IgG from the goat in the second step of the incubation process.

Sera were tested using various methods for the detection of spermatozoal antibodies. It appeared that an indirect immunofluorescence technique, a gelatine-agglutination test (Kibrick et al., 1952) and a micro-agglutination method (Franklin & Dukes, 1964) revealed different antigen-antibody systems. Sera showing a strong reaction in the gelatine-agglutination test could immobilize spermatozoa, whereas this was not the case for sera with high titres by the indirect immunofluorescence technique or for sera which were positive by the micro-agglutination test. No sera of women from infertile couples were positive by the gelatine-agglutination test, and sera from this group of women produced exclusively head-to-head agglutination in the micro-agglutination test and then only in a low titre. A few sera from men showing a reaction in the gelatine-agglutination test produced a weak tail-to-tail agglutination in the micro-agglutination test. The localization on or in the spermatozoa of the antigens with which the a...

11 HSV strains isolated from patient's material are typed by an indirect immunofluorescent technique and the results are compared to those of the “rct”-test (distinction by temperature markers). Producing both satisfactory results, the first method is superior to the second one because of its rapidity.

An antibody reacting with endocardium, vascular structures and interstitium of striated muscle (EVI antibody), by the indirect immunofluorescence technique, was recently described in Chagas' disease. In the present report the identification of tissue structures responsible for the EVI reaction was studied by means of ultrastructural immunochemical methods. Positive reactions were observed in the plasma membrane of the heart and skeletal muscle fibers and endothelial cells of blood vessels. Absorption of positive sera with T. cruzi abolished both patterns observed by electron microscopic methods. These results are in agreement with observations performed with the indirect immunofluorescence technique.

Sera from healthy people and from patients with schizophrenia were heated at 56° for 30 min and then kept at 4°. These sera were then tested for cytotoxicity and by the indirect immunofluorescence method with thymocytes, bone-marrow cells, and lymph-node cells of adult C 3 H mice. The cytotoxicity index for mouse thymocytes of sera from 60 healthy donors ranged from 0.00 to 0.4, with an average of 0.08. That of sera from 35 patients with schizophrenia ranged from 0.29 to 0.91, with an average of 0.71. The cytotoxicity index for mouse bone-marrow cells of sera from healthy donors ranged from 0.03 to 0.23, whereas that of sera from patients with schizophrenia ranged from 0.00 to 0.14. Sera of patients at dilutions of 1:8-1:32 displayed immunofluorescence of almost 100% of thymocytes and 58-78% of lymphocytes, whereas sera of healthy donors displayed none. We thus conclude that the sera of patients with schizophrenia contain antibodies against thymic antigens localized on thymocytes and thymus-derived lymphocytes.

With the direct and indirect immunofluorescence technique, deposits of immunoglobulins (predominantly IgG, less frequently IgA, IgM and IgD) and complement were demonstrated in lesional skin of patients with mycosis fungoides. The occurrence of these deposits proved to be related to the infiltrative stage of the disease. Immunoglobulin and complement deposits were not found in the premycotic stage of mycosis fungoides, but in the tumour stage a small number were demonstrable. In 2 patients with other forms of malignant reticulosis, i.e. Brill-Symmers' disease and lymphosarcoma such deposits could not be detected in the lesional skin.

A New Fluorescent Antibody Method: Mixed Antiglobulin Immunofluorescence or Labelled Antigen Indirect Immunofluorescence Staining

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.

Ontogeny and localization of the lens γ-crystallin antigen were investigated in the embryonic and post-embryonic pigeon lenses by the indirect immunofluorescence with antiserum from rabbit immunized with isolated pigeon lens γ-crystallin. The results show that γ-crystallin appears for the first time in a 4-day embryonic lens and is distributed uniformly over the entire fibre cells. In a late and also post-embryonic lens γ-crystallin was also detected in the epithelium and annular pad areas and the intensities of immunofluorescent reaction gradually increased from embryonic to post-embryonic lens.

An indirect immunofluorescence technique was applied to the typing of Herpesvirus Hominis (HVH) strains of genital and non-genital origin. The virus type was determined from aceton-fixed cells with rabbit convalescent sera and anti-rabbit conjugate. Results from kinetic neutralization confirmed the results. Cytopathic effect and plaque characteristics in Vero and chick embryo cell monolayers were evaluated for their usefullness in distinguishing type 1 strains from type 2 strains. Cytopathic effect was found to be a less reliable marker even under standardized conditions. On the other hand, the plaque size in Vero cells and the plaque forming capacity in chick embryo cell monolayers discriminated the two types.

Summary.— The basement membrane antibodies in 3 sera of patients with bullous pemphigoid were studied by means of indirect immunofluorescent staining chessboard titrations on monkey, rabbit and guinea-pig lip tissue. Comparisons of the results of these studies with those obtained in previous studies of this type with 2 other sera, revealed wide variations in the spectra of specificities of the bullous pemphigoid antibodies. Monkey oral or oesophogeal mucosa appears to be the substrate of choice for routine tests for these antibodies, although this must be regarded as an approximation of the “ideal antigen”. The implication of these findings are discussed both from the practical and theoretical points of view.

Peripheral blood lymphocytes from 38 patients with malignant lymphoma and 14 subjects with acute lymphatic leukemia were studied for relative distribution of thymus-derived (T-cells) and bone-marrow derived (B-cells). B-cells were identified using direct immunofluorescent enumeration of lymphocytes showing surface Ig, and T-cells were counted using indirect immunofluorescence and rabbit antihuman fetal thymocyte antisera. When absorbed with B-cells from patients with chronic lymphatic leukemia, the latter antisera appeared to show θ-like or T-cell specificity. Patients with lymphoma showed a significant increase (P By contrast acute lymphatic leukemia was characterized by a significant increase in peripheral blood T-cells (P

By indirect immunofluorescence, 13 different proteins were studied in 2-hour, acquired pellicles collected from four subjects over a 5-day period. The 300 specimens were coded, and the intensity of fluorescence specific for each protein was scored by two independent observers. Statistical analyses indicated that the mean score values differed significantly when a comparison was made between individual proteins or between subject-donors of the pellicles. Also, the scores for each protein were highly dependent on the subject. Despite daily fluctuations, the pellicles from each subject presented a characteristic protein profile. Frequently, high fluorescence scores were noted for proteins of generally low salivary concentration, e.g., IgM and IgG. The third component of complement was discovered in several pellicles and, by subsequent testing, in whole saliva but not in parotid fluid. Its presence in pellicles of an edentulous subject suggests that this protein may be transferred across the oral epithelium.

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.

SummaryA quick, reliable and inexpensive test is described for identifying the T or “tumor” antigen, in cells rendered neoplastic by the oncogenic DNA virus SV40. The approach is based on the feasibility of adapting the tumor imprint technique to the indirect immunofluorescence test. This method is of particular advantage when one wishes to examine leukemic or lymphosarcomatous cells, which lack the capacity to grow as monolayers in culture.