The sheepshead has two readily isolatable humoral immunoglobulins, a 16S tetrameric form and a 6S monomeric form. The 16S tetrameric form is composed of two subpopulations, one being a disulfide linked form (~700,000 daltons) and the other a noncovalently linked population of predominantly disulfide linked dimers (~350,000 daltons). The 6s immunoglobulin (~140,000 daltons) is composed of two noncovalently linked units (~70,000 daltons) each having one heavy and one light chain. The 6S immunoglobulin is antigenically deficient to the 16S immunoglobulin, this deficiency may be due to the heavy chain of the 6s protein lacking a~25,000 dalton segment present in the heavy chain of the 16S molecule. Cutaneous mucus and bile also contain immunoglobulins. The mucus contains three proteins that can be considered immunoglobulins: a 6S form which is antigenically indistinguishable from the serum 6S immunoglobulin: a ~700,000 dalton form which does not have a "dimeric" subpopulation as observed with the serum 16S protein; and a dimeric form of ~350,000 daltons. The dimeric form may have a secretory piece since the reduced mucus dimeric protein shows an additional polypeptide chain at ~95,000 daltons. All of the cutaneous mucus high molecular weight immunoglobulins have heavy and light chains identical to the serum high molecular weight immunoglobulins (~70,000 and ~25,000 daltons). Bile immunoglobulin is dimeric and composed of two noncovalently linked monomers of ~l60,000 daltons. The bile heavy chains are ~55,000 daltons; the light chains are ~25,000 daltons. The bile immunoglobulin does not appear to be a different class of protein from that of the serum or mucus immunoglobulins. In vivo administration of radiolabeled 16S and 6S serum immunoglobulins indicates that the 6S protein is not a degradation product of the 16S form. The half lives of the 16S and 6s forms are both ~16 days. Furthermore, the secretory immunoglobulins of the bile and mucus are not due to simple transudation or active transport of the predominant serum immunoglobulins. This result suggests that the secretory immunoglobulins of the sheepshead may be locally synthesized.

Transcription of the immunoglobulin heavy chain is influenced, in part, by an intronic enhancer region located 3' to the V,D, and J gene segment regions. The regulatory elements within this enhancer show high levels of sequence homology between mouse, rat, rabbit, and human, indicating their evolutionary significance. SRY, SpiB, LM02, and Rzra have been shown elsewhere to bind recognition sequences found within the evolutionarily conserved regulatory elements of this enhancer. We seek to demonstrate previously unreported individual and cooperative DNA binding activities for I these factors at regulatory' elements within the immunoglobulin 3' enhancer region. To facilitate this investigation, clones representing E2A, p65, SRY, SpiB LM02, ABF-1, and Rzra were retrieved from a plasma cell eDNA library. E2A, p65, and ABF-1 have been previously reported to bind elements within this enhancer. EMSA studies of these factors individually and in concert support previous characterization of DNA binding activities at this enhancer, and demonstrate the hereto unreported individual binding activities of Rzra, LM02, SpiB and SRY. Mixing studies with LM02 and SpiB show that when simultaneously present, these factors block the binding activity of one another. Furthermore, SpiB and LM02 are capable of blocking the in vitro DNA binding activity ofE2A and p65, presumably through the formation of a non-DNA binding complex. Mixing studies with Rzra demonstrate it to be a high affinity DNA binding factor capable of blocking p65 and E2A binding activity without relinquishing its own activity. We suspect this blocking activity to be a result of steric hinderance based on the close linear proximity of these factors recognition sequences.

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Two studies were conducted to investigate factors involved in absorption of immunoglobulins in the preruminant neonate. In the first study 26 Angus cows were fed 57% or 100% of their NRC recommended requirements for protein and energy for the last third of gestation in each of 2 years. Resulting calves were fed measured amounts of colostrum from their dam or from a cow of the reciprocal nutritional treatment group. Cows from the restricted intake treatment lost weight and body condition (P < .05). Gestation length, birth weight, calving ease, days open end weaning weight, however, were not affected by treatment. Calves born to restricted fed dame had higher serum cortisol and lower serum T₃ concentration (P < .05), but absorption of IgG was not affected. Calves fed colostrum from restricted cows did have lower serum IgG concentrations (P < 0.2); although, none of the calves were considered hypogammaglobunemic. In the second study, 2 trials were conducted to evaluate the effect of varying cortisol concentrations on Ig absorption in lambs. Treatments consisted of control (CO), high cortisol (HC), single peak of cortisol (SP) end low cortisol (LC). Lambs in trial 1 were obtained on d 136 to 138 of gestation by caesarean operation. HC and SP lambs tended to have a faster rate of Ig absorption through 24 h, but did not differ from CO lambs in serum Ig concentration by 36 h. Precocious closure to Ig absorption had occurred for LC lambs by 20 h end they had lower Ig concentrations at 36 and 48 h (P < 0.05). In trial 2, lambs were obtained on d 140 to 142 of gestation. Lambs had a shorter Ig absorptive period than trial 1 (24 vs 36 h for CO]. Premature closure for the LC lambs occurred by 16 h postpartum (P < 0.05) and they tended to have depressed Ig concentrations post 24 h. / Ph. D.

Antibodies are one of the most useful molecules with affinity of binding and specificity for in vitro and in vivo diagnosis, or for immunotherapy of human diseases. In recent years, phage-displayed antibody library has been widely adopted to select tailor-made antibodies in a fast, high-throughput mode, as an alternative of traditional hybridoma technology. Although phage display has been introduced for about 20 years, the applications and development of this technology still have a rich space to be explored. / Attempts are made in the present study to extend three applications of the phage displayed antibody library in antibody discovery and engineering. Firstly, a CDR3-randomized phage-displayed scFv library was constructed from genomic DNA of mouse. Following biopanning, anti-peptide of mas oncoprotein scFvs were isolated and identified. These results illustrate the potential use of the genomic phage-displayed library for anti-peptide antibodies selection. Secondly, we described the isolation of anti-idiotypic scFvs against a chimeric anti-CD22 mAb from an immunized phage-displayed scFv library. The isolated anti-Id scFvs were able to capture the immune response of chimeric anti-CD22 mAb with high specificity. This reagent will enhance our understanding of the therapeutic mechanism of anti-CD22 mAb in non-Hodgkin's lymphoma treatment, and may be applied to probe the pharmacokinetics, tissue distribution, and modulation of anti-CD22 mAb in vivo. / In conclusion, we have attempted various approaches to identify specific anti-peptide scFvs, anti-idiotypic scFvs and passive anti-tumor scFvs. These results extend the applications of phage display technology in antibody discovery and engineering. / Our approach enables us to isolate selective and sensitive anti-idiotypic antibodies and could be exploited for other antibodies with clinical and biological applications. Thirdly, we profile a strategy to select and identify markers on tumor cell surface using phage-displayed antibodies from mice bearing xenograft tumor. Our data imply that passive antibodies in cancer patients may be obtained from the immune repertoire of cancer patients. Besides, we found a cell surface antigen was up-regulated more than 3-fold in mas-expressing cells. We further use the targeting antibody to construct a tumor endoprotease-activated immunotoxin. / Zhao, Qi. / Adviser: Wing-Tai Cheung. / Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3499. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 227-250). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

Thesis (Ph.D.) -- University of Texas Southwestern Medical Center at Dallas, 2005. / Embargoed. Vita. Bibliography: 97-123.