RESUMEN

Carcinoembryonic antigen (CEA) was purified after perchloric acid extraction of glycoprotein from human normal and tumoral colon. Antibodies against those extracts as well as purified CEA were raised in rabbits. Sera obtained from animals immunized with tumoral extract in protracted schemes showed disperse behavior on polyacrylamide gel electrophoresis (PAGE) for proteins in the area with gamma, mobility which were markedly increased. In addition, a large increase of proteins with a mobility comparable to alpha 2 proteins was observed. The latter increase was not present in sera of rabbits immunized with normal colon extracts. Low titer antisera were obtained after immunization with either antigen as revealed by immunodiffusion. The relatively highest titers were seen against the glycoprotein fraction of tumor extracts which had previously been enriched by isoelectric focusing. Immunoelectrophoresis of tumor extracts against homologous antisera revealed bands with beta mobility due to the presence of CEA antibodies only in the sera of animals injected with tumor extracts. Antibody titers assayed by radioimmunoassay (RIA) revealed that the highest titers were obtained from tumor glycoprotein extracts was used as an immunogen. Measurements of CEA concentrations in serum of patients with low and high levels of CEA showed non significant differences by statistical analysis, when antisera obtained from different bleedings of the same animals were used. It is concluded that in order to establish a CEA RIA for clinical purposes it is necessary to purify up to the isoelectric focusing step the material to be labelled as tracer or used as standard. Antibody reagent may be obtained by immunizing schemes or protracted schemes using crude perchloric acid extracts of colon tumor.

Asunto(s)

RESUMEN

Carcinoembryonic antigen (CEA) was purified after perchloric acid extraction of glycoprotein from human normal and tumoral colon. Antibodies against those extracts as well as purified CEA were raised in rabbits. Sera obtained from animals immunized with tumoral extract in protracted schemes showed disperse behavior on polyacrylamide gel electrophoresis (PAGE) for proteins in the area with gamma, mobility which were markedly increased. In addition, a large increase of proteins with a mobility comparable to alpha 2 proteins was observed. The latter increase was not present in sera of rabbits immunized with normal colon extracts. Low titer antisera were obtained after immunization with either antigen as revealed by immunodiffusion. The relatively highest titers were seen against the glycoprotein fraction of tumor extracts which had previously been enriched by isoelectric focusing. Immunoelectrophoresis of tumor extracts against homologous antisera revealed bands with beta mobility due to the presence of CEA antibodies only in the sera of animals injected with tumor extracts. Antibody titers assayed by radioimmunoassay (RIA) revealed that the highest titers were obtained from tumor glycoprotein extracts was used as an immunogen. Measurements of CEA concentrations in serum of patients with low and high levels of CEA showed non significant differences by statistical analysis, when antisera obtained from different bleedings of the same animals were used. It is concluded that in order to establish a CEA RIA for clinical purposes it is necessary to purify up to the isoelectric focusing step the material to be labelled as tracer or used as standard. Antibody reagent may be obtained by immunizing schemes or protracted schemes using crude perchloric acid extracts of colon tumor.

RESUMEN

The sera of three groups (I, II and III) of cattle vaccinated every three months with trivalent hydroxysaponinated commercial vaccine against aphthovirus were studied. The only difference between groups I and II was that the former received a revaccination on day 17 after the initial immunization. Groups I and II included sera from animals three months old born from vaccinated mothers. Group III consisted of the sera of adult animals (the mothers of animals in groups I and II). The animals from the three groups were bled monthly during one year. The studies were performed with pooled sera from each group. The presence of protective and neutralizing antibodies was investigated in the gammaglobulin fractions which were then separated into subclasses, by chromatography on DE-cellulose columns, in order to study their biological activity. The immunization of cattle 3 months old with commercial vaccine against aphthovirus resulted in weak primary humoral response; neutralizing antibodies could not be detected. When the animals were restimulated three weeks after the first immunization, neutralizing antibodies appeared although the response did not persist. Nevertheless, five months after the experiment was started both groups I and II showed neutralizing antibodies. (Fig. 1, 2, 3). Persistent immunity to the three virus subtypes was acquired by animals of groups I and II but not before nine months. The kinetics of protective antibodies was similar to that of neutralizing antibodies, but with higher titers. Some bleedings that did not show neutralizing activity, did show significant protective activity (Figs. 4, 5). The investigation of the neutralizing activity of the gammaglobulin subclasses obtained by chromatography revealed that there was not one single subclass responsible for this activity, but that several subclasses were involved. The gammaglobulin subclasses were analyzed by immunoelectrophoresis; proteins with alpha 2 mobility appeared, coincident with early bleedings of high neutralizing titers, although these proteins did not present neutralizing activity (Tables 1, 2). The protective and neutralizing activity was not correlated with the protein concentration of the fractions so that the increase observed may be due to a qualitative change in the antibodies.

Asunto(s)

RESUMEN

The sera of three groups (I, II and III) of cattle vaccinated every three months with trivalent hydroxysaponinated commercial vaccine against aphthovirus were studied. The only difference between groups I and II was that the former received a revaccination on day 17 after the initial immunization. Groups I and II included sera from animals three months old born from vaccinated mothers. Group III consisted of the sera of adult animals (the mothers of animals in groups I and II). The animals from the three groups were bled monthly during one year. The studies were performed with pooled sera from each group. The presence of protective and neutralizing antibodies was investigated in the gammaglobulin fractions which were then separated into subclasses, by chromatography on DE-cellulose columns, in order to study their biological activity. The immunization of cattle 3 months old with commercial vaccine against aphthovirus resulted in weak primary humoral response; neutralizing antibodies could not be detected. When the animals were restimulated three weeks after the first immunization, neutralizing antibodies appeared although the response did not persist. Nevertheless, five months after the experiment was started both groups I and II showed neutralizing antibodies. (Fig. 1, 2, 3). Persistent immunity to the three virus subtypes was acquired by animals of groups I and II but not before nine months. The kinetics of protective antibodies was similar to that of neutralizing antibodies, but with higher titers. Some bleedings that did not show neutralizing activity, did show significant protective activity (Figs. 4, 5). The investigation of the neutralizing activity of the gammaglobulin subclasses obtained by chromatography revealed that there was not one single subclass responsible for this activity, but that several subclasses were involved. The gammaglobulin subclasses were analyzed by immunoelectrophoresis; proteins with alpha 2 mobility appeared, coincident with early bleedings of high neutralizing titers, although these proteins did not present neutralizing activity (Tables 1, 2). The protective and neutralizing activity was not correlated with the protein concentration of the fractions so that the increase observed may be due to a qualitative change in the antibodies.

RESUMEN

The sera of three groups (I, II and III) of cattle vaccinated every three months with trivalent hydroxysaponinated commercial vaccine against aphthovirus were studied. The only difference between groups I and II was that the former received a revaccination on day 17 after the initial immunization. Groups I and II included sera from animals three months old born from vaccinated mothers. Group III consisted of the sera of adult animals (the mothers of animals in groups I and II). The animals from the three groups were bled monthly during one year. The studies were performed with pooled sera from each group. The presence of protective and neutralizing antibodies was investigated in the gammaglobulin fractions which were then separated into subclasses, by chromatography on DE-cellulose columns, in order to study their biological activity. The immunization of cattle 3 months old with commercial vaccine against aphthovirus resulted in weak primary humoral response; neutralizing antibodies could not be detected. When the animals were restimulated three weeks after the first immunization, neutralizing antibodies appeared although the response did not persist. Nevertheless, five months after the experiment was started both groups I and II showed neutralizing antibodies. (Fig. 1, 2, 3). Persistent immunity to the three virus subtypes was acquired by animals of groups I and II but not before nine months. The kinetics of protective antibodies was similar to that of neutralizing antibodies, but with higher titers. Some bleedings that did not show neutralizing activity, did show significant protective activity (Figs. 4, 5). The investigation of the neutralizing activity of the gammaglobulin subclasses obtained by chromatography revealed that there was not one single subclass responsible for this activity, but that several subclasses were involved. The gammaglobulin subclasses were analyzed by immunoelectrophoresis; proteins with alpha 2 mobility appeared, coincident with early bleedings of high neutralizing titers, although these proteins did not present neutralizing activity (Tables 1, 2). The protective and neutralizing activity was not correlated with the protein concentration of the fractions so that the increase observed may be due to a qualitative change in the antibodies.