Reactions of human and hog intrinsic factors with type I antibody to intrinsic factor

A simple and rapid small-scale gel filtration method was applied in studies of type I antibody to intrinsic factor using radioactive vitamin B₁₂ of high specific activity and purified human and hog intrinsic factor preparations, taking into account the unsaturated B₁₂-binding capacity of the individual pernicious anaemia sera. This procedure allows the use of small amounts of reagents. Evidence was obtained for a close antigenic similarity of determinants of human and hog intrinsic factor. The use of purified intrinsic-factor preparations is important.     

Studies on rabbit secretory and serum antibodies to hog intrinsic factor

The response of the systemic and secretory immune systems to hog intrinsic factor (IF) was studied in rabbits. Following immunization, pregnant rabbits produced both serum and colostral antibodies to IF. Blocking and binding antibodies to IF were consistently found in sera. The colostrum of the four animals studied also contained blocking antibodies, but colostral-binding antibodies to IF were detected in only two rabbits. Blocking antibody in serum was restricted to the IgG fraction, whereas this activity was present in both the IgG and secretory IgA immunoglobulins of colostrum. Binding antibody to IF was also found within the IgG and secretory IgA of colostrum; in serum, however, the binding activity was seen in all three immunoglobulin classes. These data indicate that the systemic and secretory immune systems of the rabbit recognize the two major antigenic sites on IF and produce specific antibodies against these determinants.     

Studies of the antigenic structure of Coxiella burnetii

The antigenic structure of Coxiella burnetii (C.b.) was studied by absorption of human and animal immune sera with C.b. organisms in the natural phase II (NPh II) or with artificial phase II (ArPh II) organisms prepared by their treatment with KIO4. It was found that immune sera absorbed with one type of phase II organisms still reacted with the antigen of another type of phase II organisms as demonstrated in both microagglutination (MA) and complement-fixation (CF) tests.     

Experimental humoral and cellular immunity to hog intrinsic factor

Humoral and cellular immunity to hog intrinsic factor (HIF) was studied in rabbits immunized with varying doses of HIF or HIF complexed to vitamin B₁₂. Animals immunized with large doses of HIF (1 mg) consistently produced high titres of blocking and binding antibodies to IF. At low dose immunization (10 μg), the humoral response was obviously blunted, with animals forming significantly reduced titres of antibodies to IF; several rabbits in this group made only binding antibodies and one rabbit produced neither blocking nor binding antibodies. No difference in humoral responsiveness was noted between those animals who received HIF or an equivalent amount of HIF complexed to vitamin B₁₂. By contrast, cellular immunity as measured by inhibition of leucocyte migration was readily induced to a similar degree in both high and low dose animals, including the rabbit which had no detectable antibodies to IF. When an intermediate dose of HIF (50 μg) was used as the immunogen, four rabbits gave positive leucocyte migration tests only with HIF—vitamin B₁₂ complex, suggesting that the interaction of HIF with vitamin B₁₂ may enhance its antigenicity in vitro. These data demonstrate that cellular and humoral immunity to HIF can be induced and partially dissociated from each other by varying the dose of immunogen. Although the mechanism responsible for this dissociation is not clear, one explanation would be differing sensitivities of lymphocyte-populations to different doses of antigen.     

Studies on the glycosylation of flavivirus E proteins and the role of carbohydrate in antigenic structure

The glycosylation pattern of several flavivirus E proteins as well as the role of carbohydrate in biological functions and the antigenic structure of tick-borne encephalitis (TBE) virus were investigated by the use of specific endoglycosidases. Endoglycosidase F digestion revealed the presence of a single asparagine-linked oligosaccharide side chain in TBE virus (Western and Far Eastern subtype), Louping III virus, Murray Valley encephalitis virus, and Rocio virus. Consistent with published sequence data, the E protein of West Nile virus apparently is not glycosylated at all. Evidence derived from digestion experiments using endoglycosidase H indicates that the tick-borne viruses contain high-mannose type N-linked oligosaccharide side chains, whereas that of the mosquito-borne Murray Valley encephalitis virus and Rocio virus is endoglycosidase H resistant. Complete deglycosylation of TBE virus by endoglycosidase F did not impair infectivity and HA activity. Carbohydrate does not seem to play a major role in the antigenic structure of the TBE virus glycoprotein since the reactivity of the native virus and the deglycosylated virus was identical when analyzed with monoclonal as well as polyclonal immune sera.     

Immunogenicity and specificity of collagen: VIII. Studies on the antigenic structure of soluble fish collagen

In rabbit antisera to acid-soluble carp collagen two antibody fractions differing in specificity were demonstrated by passive haemagglutination. The antibody fractions were separable from each other on an immunoadsorbent prepared from rabbit collagen. One reacted exclusively with carp collagen, the other reacted additionally with various mammalian collagens including that from rabbit. The results were interpreted as evidence for the presence of at least two different antigenic determinants on carp collagen resembling former results for calf collagen.     

Study of the antigenic structure of human serum albumin with monoclonal antibodies

Analysis of the antigenic structure of human serum albumin was undertaken using monoclonal antibodies. Nineteen antibodies were prepared and their specificities were studied using fragments which encompass the whole sequence of the albumin molecule. These antibodies recognized 13 different epitopes which are different from the one previously identified with two other monoclonal antibodies [Doyen et al., Immun. Lett. 3, 365-370 (1981)]. Among those 13 different epitopes, six were overlapping. Four epitopes were located on the N-terminal half of the albumin molecule. One of these required integrity of methionine 87 and the other three were overlapping and located around methionine 123. Eight epitopes were located on the C-terminal half of the albumin. Two of them were within the sequence, 330-422 and 299-496 respectively; the other six appeared to be topographic determinants which were altered or lost in the albumin fragments. A last epitope could not be located on any region of albumin. Four monoclonal antibodies directed against a given portion of the albumin molecule reacted slightly with another part of albumin, thus confirming the existence of an intramolecular cross-reactivity between the different domains of human albumin.     

Mitochondrial antigenic structure and enzyme activity in ageing human diploid fibroblasts

It has been proposed that cellular ageing may be caused by loss of mitochondrial function due to the action of free radicals. To investigate this hypothesis, antigenic structures of the mitochondrial inner membrane/matrix and of the outer mitochondrial membrane of human diploid fibroblasts were monitored by immunoblotting at four stages during cellular lifespan in vitro. At the same time, specific activities of the enzymes oligomycin-sensitive ATPase (O-S ATPase), malate dehydrogenase (MDH) and glutamate dehydrogenase (GDH) were assayed to assess the functional capacity of cellular oxidative phosphorylation and of the tricarboxylic acid cycle. No changes were found with ageing in inner mitochondrial membrane-associated matrix components, or in the activities of O-S ATPase and MDH. However GDH activity increased significantly with ageing in vitro, possibly indicating greater amino acid utilization for energy production in older cells. There was loss of an outer mitochondrial membrane antigen, of approximate molecular weight 60 kilodaltons (kDa), in the oldest cells tested, which may influence outer membrane transport capacity late in the cellular lifespan. Overall, the results fail to provide support for the hypothesis that ageing primarily results from free radical-induced impairment of mitochondrial function.