Immunochemical study of the specific antigens of human amniotic fluid]

Four specific antigens (trophoblastic beta 1-globulin, placental lactogen, alpha 1- and alpha 2-globulins of human placenta) were identified using antisera to the native amniotic fluid. Five antigens with the mobility of prealbumins, alpha 1-globulins, alpha 2-globulins and beta 2-globulins which bear no resemblance with the previously studied antigens were identified using antisera to the acid fraction of the amniotic fluid. Both the prealbumins and alpha 2-globulin were found in the blood serum of foetuses of different age and of newborn infants; these proteins were absent from the blood serum of pregnant women and donors. They received the names of embryonic prealbumine 1, embryonic prealbumine 2 and embryonic alpha 2-globulin. The protein with the mobility of alpha 1-globulins was found in the amniotic fluid of foetuses and in the blood serum of pregnant women only and received the name of amniotic alpha 1-globulin. The concentration of the antigens in question was studied in the developing foetuses and in the blood serum of pregnant women at different stages of pregnancy.     

A blocking factor in normal animal sera inhibiting immunologic reactions in vitro]

An extremely thermostable and resistant to some chemical substances factor was found in the sera of Wistar and noninbred rats of different age, male and female. This factor inhibited the action of normal and immune antibodies to rat erythrocytes and rat serum antigens. The blocking factor was relatively specific and more effective in respect to the normal antibodies than to the immune ones. The activity of the blocking factor was connected with the alpha1-globulin.     

Immunochemical determination of placenta-specific and interorganic antigens in placental extract and blood serum in pregnant rats

It has been demonstrated that placenta extract of rats contains up to 14 antigens. Moreover, 11 of them are interorgan proteins of wide and limited specificity, two antigens (alpha 1- and alpha 2-globulins) are attributed to acute-phase proteins typical for pregnancy. beta 1-Globulin is a specific protein of rat placenta. The content of these antigens in blood serum increases with pregnancy and reaches a maximum toward the delivery; 3-4 days after delivery beta 1-globulin disappears completely from maternal blood, whereas the concentration of acute-phase proteins drops to the initial level.     

Immunoenzyme analysis of the fertility alpha 2-microglobulin in the blood serum of men and women

It has been found that fertility alpha 2-microglobulin content in male and female serum does not exceed 20 ng/ml and 40 ng/ml, respectively. A high level of fertility alpha 2-microglobulin was found in the serum in early pregnancy, with its concentration decreased by parturition.     

Tissue-specific retinal proteins in vertebrate evolution

Studies have been made on water soluble antigens of the retina from man and some animals. In the bovine retina, immunochemical analysis reveals, apart from antigens with a broad and narrow interorganic specificity, organospecific alpha 1- and rho-globulins. Immunochemically, the bovine alpha 1-globulin is partially identical with the same protein of the human retina and completely identical to retinal antigens from cattle; rho-globulin is characterized as an interspecific antigen in man and mammals. Molecules of organospecific alpha 1-globulins from the retina of man and some animals (sheep, camel, horse, cow, pig) do not contain the determinants related to the retinal antigens from fishes, reptiles and birds. In human and mammalian retina, acid neurospecific alpha 1-glycoprotein was found which is topical of the cerebral tissue. Organospecific alpha 1-globulin of the bovine retina is located in the pigment epithelium, in the zone of outer and inner photoreceptor segments; organospecific rho-globulin is distributed in the outer synaptic layer of the retina.     

Isolation and characteristics of a rabbit β2-microglobulin: Comparison with human β2-microglobulin

A low molecular weight β₂-globulin has been isolated from urine of rabbits treated with sodium chromate. Several findings indicate that this protein is the rabbit counterpart of human β₂-microglobulin which is known to occur linked to the major histocompatibility (HL-A) antigens of man. The rabbit β₂-globulin and human β₂-microglobulin are very similar in amino acid composition, molecular size, molecular weight, electrophoretic mobility, and also in the presence of apparently analogous disulfide loops. More than half of the amino acids in the two proteins seem to be present in identical numbers. The urinary excretion of both proteins is increased by agents which induce renal tubular damage.     

Immunochemical study of the CBA mouse kidney]

Immunoelectrophoresis and agar precipitation reaction allowed one to reveal 7 antigens in CBA mouse kidney. One of them, with a relative electrophoretic mobility--0.35, is specific for the kidney. The remaining renal antigens are common to the kidney and other organs and tissues of mice. The antigens with mobility in the albumin and alpha-globulin zones proved to be serum ones. An antigen with a mobility approximating 0 is specific for the kidney and liver. An antigen with a mobility in the alpha 2-globulin zone does not seem likely to be homogeneous and, apart from broad inter-organ specificity, bears the greatest resemblance to the lungs antigen.     

The influence of the thermal factor on the antigenic properties of skin]

Results of immunochemical studies of normal and thermally-treated human skin in vitro showed at least 2 of 4-5 organospecific dermal antigens to be thermostable-they withstood heating at 100 degrees C for 3 minutes; 2 or 3 antigens were thermolabile. The thermostable antigens possessed electrophoretic migration in the field of alpha1- and gamma-globulins. The burn eschar was found to retain one of the two thermostable oranospecific antigens together with the loss of thermolabile antigen.     

Proteolysis of the heavy chain of major histocompatibility complex class I antigens by complement component C1s.

The major histocompatibility complex (MHC) class I antigens contain a light chain, beta 2-microglobulin, non-covalently associated to the transmembrane heavy alpha-chain carrying the allotypic determinants. Since the C1q complement component is known to associate with beta 2-microglobulin, and we recently found that activated C1s complement was capable of cleaving beta 2-microglobulin, we decided to investigate the proteolytic activity of C1 complement towards the heavy chain of class I antigens. Our results demonstrate that human C1s complement cleaves the heavy chain of human class I antigens into at least two fragments, with apparent molecular weights of 22,000 and 24,000 g/mol on sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), under both reducing and non-reducing conditions. The cleavage of the heavy chain is inhibited by the presence of C1 esterase inhibitor. The molecular weights of the fragments are in agreement with the cleavage located in the area between the disulphide loops of the alpha 2-and alpha 3-domains of the heavy chain. In addition human C1s complement is able to cleave H-2 antigens from mouse in a similar fashion but not rat MHC class I antigen or mouse MHC class II antigen (I-Ad). Mouse MHC class I antigen-specific determinants could also be detected in supernatant from mouse spleen cells incubated with C1r and C1s. These results indicate the presence in the body fluids of a non-membrane-bound soluble form of the alpha 1-and alpha 2-domains which represent the binding site for antigenic peptides.     

Complement component C1r mediated cleavage of the heavy chain of the major histocompatibility class I antigens.

Apart from cleaving C1s, we demonstrate for the first time that: 1) at concentrations found in serum, the activated forms of the complement components C1r in addition to C1s can cleave the heavy chain of MHC class I antigens, 2) the cleavage by C1r and C1s is seemingly dependent upon a native configuration of the MHC class I antigen, since heat denaturation of the HLA antigens reduce the cleavage. The proteolytic fragments following C1 cleavage were characterized by precipitation with Con A-Sepharose, anti-MHC class I and anti-beta 2-microglobulin antibodies. The proteolysis of the alpha-chain of MHC class I was shown to take place between the alpha 2- and alpha 3- domains as estimated by the Con A-Sepharose precipitation pattern on SDS-PAGE. The alpha 1/alpha 2 fragment was still shown to interact with beta 2-microglobulin as shown by immunoprecipitation.     

Immunochemical analysis of the interaction of the fertility alpha 2-microglobulin with steroid sex hormones

The interaction between the fertility alpha 2-microglobulin and steroid sex hormones (estrone, estriol, estradiol, progesterone, testosterone) was studied by the use of cross immunoelectrophoresis with intermediate gels. alpha 2-microglobulin was shown to bind to the above hormones, its affinity to testosterone being the highest. The ability of alpha 2-microglobulin to bind to steroid hormones can be used for its isolation by affinity chromatography with immobilized steroid hormones.     

Identification, isolation and physico-chemical properties of neurospecific alpha2-globulin]

In the immunization process of rabbits with the protein fraction of the water-salt extract from human brain partially and concentrated at 500-600 times was received antiserum revealed brain specific alpha 2-globulin that is not identical to the known cytoplasmatic brain specific antigens. This antigen has got electrophoretic mobility of alpha 2-globulins, molecular weight 90 +/- 10 kD and isoelectric point 4.1-5.4. Develop the procedure for purification of this antigen on the basis of the combination ion change, affinity, hydrophobic chromatography gel-filtration and isochromatofocusing.     

Thermostable antigens of malignant tumors and normal tissues of experimental animals]

Gel precipitation and immunoelectrophoresis tests tests demonstrated the presence of thermostable antigens in dimethylbenzanthracene-induced malignant tumours of the muscle tissues of Wistar rats. These antigens were relatively tissue-specific for the tumours, amniotic fluid and the uterus on the one hand, and for the lung, spleen, and the serum on the other hand. The thermostable antigens of the tumours and the amniotic fluid had the electrophoretic mobility in the zone of alpha and alpha1-globulins.     

Immunochemical identification and physico-chemical characteristics of specific alpha 2 globulin of human granulocytes]

The antigen which has alpha 2-globulin mobility was identified immunochemically in leukolysate and pus extract. This antigen was localized in polymorphonuclear leukocytes by the immunofluorescence technique in the blood of healthy donors. The alpha 2-globulin of granulocytes (alpha 2GG) is not identical to lactoferrin, lysozyme, granulocyte elastase, fibronectin, fetal hemoglobin, amyloid P-component of the serum. The molecular mass of alpha 2GG is equal to 50 +/- 8 Kd. in the pus extract. The biological role of alpha 2GG is unknown.     

Preliminary studies on allotypes in cattle1

This paper presents evidence of five isoimmune sera, which identified five antigens of cattle blood serum. These antigens were examined by means of staining, immuno-electrophoresis and Sephadex G 200 column chromatography. It was established that the BA-2, BA-3, BA-4 and BA-4′ antigens were proteins, whilst the BA-1 antigen was a lipoprotein. The BA-3 antigen migrated in the γ-globulin position; the remaining four antigens migrated in the α-globulin region. The antigens under examination were already present in the blood serum during the first few days of life and appeared to be of a stable character. The results of genetic analysis suggest that these antigens were transmitted according to Mendelism.     

The effect of alpha2-macroglobulin from bovine serum on bovine alpha-chymotrypsin.

Bovine alpha2-globulin contains a protein which increases the activity of bovine alpha-chymotrypsin against synthetic substrates. The active protein fraction migrates slowly on polyacrylamide gel electrophoresis, so it was named slow alpha2-globulin (Salpha2). The fraction was isolated from bovine serum and purified. Its sedimentation constant S20 was 18.5 S. It was thus identified with the alpha2-macroglobulin (alpha2M). By kinetic studies, the dissociation constant of the alpha-chymotrypsin-alpha2 M complex was calculated to be of the order of 10(-7) l/mol. The purified alpha2 M was shown to bind alpha-chymotrypsin at a definite rate. If the binding ratio was assumed to be 1:2, the molecular weight was calculated to be about 8 X 10(5).     

Characteristics of heterologous antisera against stromal fibroblasts]

As shown with the use of heterologous rabbit antifibroblast sera (AFS) to stromal mechanocytes of guinea pig, the media from the cultured bone marrow, spleen, and thymus stromal fibroblasts contained specific trypsinoresistant fibroblast protein (AG-1), which was also present in the normal blood serum and on the surface of stromal fibroblasts. AG-1 was insensitive to collagenase effect and apparently differed from the chief surface protein of fibroblasts (CSP). AG-1 is referred to gamma1-globulin, and is probably a new specific surface fibroblast protein. AFS contained also antibodies to the nonspecific protein of fibroblasts (AG-2), alpha1,-globulin related to AG1.     

Structural relationship between alpha 1-microglobulin from man, guinea-pig, rat and rabbit

Rabbit alpha 1-microglobulin was purified from the urine of sodium-chromate-treated animals by the use of gel chromatography on Sephadex G-100, affinity chromatography on concanavalin-A--Sepharose and ion-exchange chromatography on DEAE-Sephadex. Rabbit alpha 1-microglobulin had a molecular mass of 25.6 kDa on SDS/polyacrylamide gel electrophoresis. Alpha 1-microglobulin has previously been purified from the urine of humans, guinea-pigs and rats by similar methods, and the molecular masses of the four homologues were compared by SDS/polyacrylamide gel electrophoresis and gel chromatography in a denaturing medium. By these two methods the human homologue was 6 kDa and 3 kDa larger, respectively, than the other three proteins. Endoglycosidase F digestion of alpha 1-microglobulin, followed by SDS/polyacrylamide gel electrophoresis, revealed three protein bands in the human alpha 1-microglobulin sample, and only two bands in guinea-pig, rat and rabbit alpha 1-microglobulin, with a gap between each band of 2.6--2.9 kDa. The amino-terminal amino acid sequences of the four homologues were determined and between 72% and 81% homology was seen. The five amino-terminal amino acids present in the other species were missing in guinea-pig alpha 1-microglobulin. Our results indicate that human alpha 1-microglobulin is substituted with two N-linked oligosaccharides, while only one is attached to each of the other alpha 1-microglobulins, and that the extra glycosylamine-linked oligosaccharide in the human protein is attached to asparagine in position 17. Finally it is shown that all four homologues inhibit antigen stimulation of human lymphocytes, a finding which is consistent with our previous suggestion that the N-linked oligosaccharides carry the immunosuppressive activity of alpha 1-microglobulin.     

Alpha chain of HLA-DR transplantation antigens is a member of the same protein superfamily as the immunoglobulins

Four cDNA clones, pDR-α-1, pDR-α-2, pDR-α-3 and pDR-α-4, corresponding to the alpha chain of HLADR antigens, have been sequenced. Restriction maps and sequences suggest that all clones are identical apart from a single-base substitution present in pDR-α-1. Amino acid sequence data, together with the nucleotide sequence data, allowed the complete amino acid sequence to be predicted. The alpha chain is composed of 229 amino acids, of which 191 are exposed on the outside of the plasma membrane. The membrane-embedded portion of the chain consists of 23 hydrophobic amino acids. The succeeding 15 amino acids form the cytoplasmically localized hydrophilic tail. The extracellular portion, with carbohydrate moieties linked to Asn⁷⁸ and Asn¹¹⁸, seems to be organized into two domains. The second domain, which contains the only disulfide bond of the alpha chain, displays amino acid sequence homology to immunoglobulin constant regions, to the second domain of the beta chain of a class II antigen, to the third domain of heavy chains of class I antigens and to β2-microglobulin. Thus the subunits of immunoglobulins, class I antigens and class II antigens are related evolutionarily.     

Critical evaluation of α1- and β2-microglobulins in urine as markers of cadmium-induced tubular dysfunction

The purpose of the study was to examine the validity of alpha1-microglobulin (alpha1-MG) in comparison with popularly used beta2-microglobulin (beta2-MG). A database was revisited to select ca. 7,500 spot urine samples (of adequate urine density) from non-pregnant, non-lactating and never-smoking adult women. The validity of the MGs was examined in terms of stability of the MG-uria prevalence in urine samples of various creatinine (CR or cr) concentration or specific gravity (SG or sg). Comparisons were made for MGs as observed (e.g., alpha1-MGob), as corrected for CR (e.g., alpha1-MGcr) and as corrected for SG of 1.016 (e.g., alpha1-MGsg). A cut-off value of 5.7 mg/g cr (or mg/l) for alpha1-MG was deduced from a cut-off value of 400 microg/g cr (or mcirog/l) for beta2-MG, because the correlation between alpha1-MGcr and beta2-MGcr was statistically significant. The prevalence of a 1-MGsg-uria was essentially unchanged (i.e., from a low of 13.6% to a high of 17.0%, or 1.2 times) except for in very dense or very thin urine samples, in contrast, beta2-MGcr-uria showed a substantial increase (from 0.0% to 2.8% with an infinite rate) as a reverse function of a decrease in CR in urine. The prevalence of uncorrected markers, i.e., alpha1-MGob-uria and beta2-MGob-uria, showed even greater CR- or SG-dependent changes. Thus, it appeared prudent to consider a alpha-MGsg rather than beta2-MGcr as a marker of tubular dysfunction among a general population with various urine density.