An improved sample applicator for column chromatography.
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Chromatography↗
Biomedical subjects
L S Rodkey
Publications and source records attributed to L S Rodkey.
Antibody synthesis induced by endogenous internal images.
In this study, immunization with a vaccine consisting of multiple F(ab')2 fragments of affinity-purified antitetanus toxoid antibodies covalently bound to a carrier protein successfully induced antitetanus toxoid antibodies. Further studies showed that this vaccine preparation contained no biologically detectable tetanus antigen. The induced antitetanus antibody (Ab1') titer was higher than the titer of antibodies binding control antigens. The immunizing F(ab')2 preparation did not elicit a secondary antitetanus response from mice primed with tetanus toxoid and, hence, appeared free of tetanus epitopes. The specificity of Ab1' was established by absorption and inhibition with antigen. Immunization with antitetanus F(ab')2 (Ab1') fragments appears to have elicited naturally occurring autologous antitetanus toxoid antibody (Ab1') through an idiotypic pathway. As predicted by network theory, anti-idiotype (Ab2) and antitetanus (Ab1') cycled reciprocally. Clonotypic characterization of Ab1' using isoelectric focusing and affinity immunoblotting showed increases in Ab1' titer to be the result of increased synthesis by limited subsets of antitetanus toxoid B-cell clones and not increased synthesis by multiple clones, as is characteristic of antigen-driven Ab1 responses. Many Ab1 and Ab1' clonotypes had identical pIs, suggesting that they either share V region genes or are the product of the same B-cell clones. These findings indicate that immunization with polyclonal multivalent Ab1 preparations can trigger active synthesis of antibodies with the same specificity. The results provide further evidence for naturally occurring idiotypic cascades that could be exploited for studies of catalytic antibodies.
Animals↗
Comparison of the immune response to Ars-BGG in germfree or conventional piglets.
Neonatal germfree (GF) colostrum-deprived and conventional (CV) colostrum-fed piglets were immunized IP with p-azo-phenyl-arsonate-bovine gamma globulin (Ars-BGG) in Freund's adjuvant to study the development of the immune response in the absence or presence of maternal antibodies and environmental antigens. Overall, the immune response varied greatly within each group but did not differ in GF from CV piglets statistically. Affinity immunoblot analysis suggested that anti-Ars antibody was more restricted in GF than CV piglets and clonotype shifts occurred more in GF than CV piglets after each antigenic stimulation. In contrast, the clonotype pattern of the anti-BGG antibody was similarly heterogeneous in the two groups. Based on the affinity immunoblot data the antibodies generated to the Ars-haptenic group in CV piglets are more heterogeneous than GF piglets and suggest that clonotype generation is influenced by maternal antibodies and environmental antigens.
Animals↗
Ultrasound-guided cardiac blood sampling of the rabbit fetus.
The object of the study was to determine whether cardiac sampling of the rabbit fetus could be successfully accomplished with minimal procedure-related loss. Pregnant rabbit dams were randomized to undergo ultrasound-guided fetal cardiac sampling in either the left or right uterine horn at 27 days of gestation; cesarean delivery was performed the following day. Liveborn pups from unsampled uterine horns underwent cardiac puncture immediately after birth. Fetal hematologic parameters were then compared to neonatal parameters. The acute fetal mortality from the sampled uterine horns was similar to that of the unsampled horns (3.6 vs. 4.5%). Fetal hematologic values were significantly higher than neonatal values with the exception of the reticulocyte count. This rabbit model offers a new approach for the evaluation of novel treatment modalities for hemolytic disease of the human fetus.
Animals↗
DNA hydrolysis by monoclonal autoantibody BV 04-01.
Monoclonal anti-DNA autoantibody BV 04-01 catalyzed hydrolysis of DNA in the presence of Mg2+. Catalysis was associated with BV 04-01 IgG, Fab, and single-chain-antibody (SCA) proteins. Cleavage of both ss and dsDNA was observed with efficient hydrolysis of the C-rich region of A7C7ATATAGCGCGT2, as well as a preference for cleaving within CG-rich regions of dsDNA. Data on specificity of ssDNA hydrolysis and kinetic data obtained from wild-type SCA, and two SCA mutants were used to model the catalytically active antibody site using the previously resolved X-ray structure of BV 04-01. The resulting model suggested that the target phosphodiester bond is activated by induction of conformational strain. In addition, the antibody-DNA complex contained a Mg2+ coordination site composed of the L32Tyr and L27dHis side chains and a DNA 3'-phosphodiester group. Induction of strain along with the metal coordination could be part of the mechanism by which this antibody catalyzes DNA hydrolysis. Sequence data for BV 04-01 V(H) and V(L) genes suggested that the proposed catalytic-antibody active site was germline-encoded. This observation suggests that catalytic activity might represent an important-rarely examined-function for some antibody molecules.
Animals↗
Electrodissociation of high molecular weight complexes of interferon alpha during recycling isoelectric focusing.
Natural human interferon alpha has been separated by selective ultrafiltration into low molecular weight components and the molecules exceeding 100K daltons. Interferon associated with a higher molecular weight fraction showed partial pH sensitivity and resisted dissociation after treatment with urea, mercaptoethanol, sodium chloride or significant changes in pH. However, interferon activity was released from high molecular weight components during recycling isoelectric focusing. Electrodissociation was carried out in 1% ampholytes for 574 watt-hours. The interferon activity was concentrated in a pH range of 6-6.5, whereas, the majority of proteins were generally found in a more acidic position. The dissociated interferon was neutralized by polyclonal antibody to human interferon alpha (IFN alpha) and showed no presence of pH labile form. A pH sensitivity of high molecular weight interferon (HMW-IFN) may reflect an aggregation phenomenon rather than intrinsic structural differences.
Cell Line↗