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A J Guidry

Publications and source records attributed to A J Guidry.

At least 55 records · Page 3Linked to original sources
PubMed1986

Radioimmunoassay for measurement of bovine alpha-lactalbumin in serum, milk and tissue culture media.

A sensitive, specific radioimmunoassay for bovine alpha-lactalbumin (alpha-la) suitable for measurement of serum, tissue culture media and milk was developed. alpha-La was not detected in serum from prepubertal male or female cattle, but was detected as early as 60 d of gestation in nulliparous Holstein heifers, the level being greatly increased during the last one third of gestation. Serum from cross bred beef heifers contained less alpha-la and it was not detected until late gestation. Concentrations of alpha-la in serum samples from pregnant multiparous Holstein cows decreased at drying-off and subsequently increased just before parturition. Secretion of alpha-la by mammary tissue explants from steroid-primed prepubertal Holstein heifers was induced by the addition of bovine prolactin, ovine prolactin or human growth hormone to tissue culture media.

Anestrus↗
PubMed1986

Immunoglobulin isotype concentrations in milk as affected by stage of lactation and parity.

Quarter milk samples were collected during lactation and analyzed for immunoglobulins from 32 first and 16 third lactation Holstein cows that were equally distributed progeny of sires selected for yield or merit. Variance component estimates indicated large variation for cow, quarter within cow, and stage of lactation. All immunoglobulin isotype concentrations were lowest at wk 21. Significant differences between first and third lactation cows were observed for milk immunoglobulin A (.012 and .018 mg/ml) and immunoglobulin M (.074 and .101 mg/ml). Quarters with cell counts above 1 X 10(6) cells/ml had higher concentrations of immunoglobulin G1, (.619 vs. .394 mg/ml) immunoglobulin G2 (.103 vs. .063 mg/ml), immunoglobulin A (.028 vs. .014 mg/ml), and immunoglobulin M (.117 vs. .087 mg/ml). There were no significant differences in immunoglobulin isotype concentrations due to genetic group (yield or merit), suggesting that selection for milk production has no significant effect on immunoglobulin concentration in milk. Degree of parity, however, must be considered when comparing immunoglobulin A and immunoglobulin M concentrations in milk.

Animals↗
PubMed1985

Quantitation of bovine beta 2-microglobulin: occurrence in body fluids, on milk fat globules and origin in milk.

The concns of bovine beta 2-microglobulin (beta 2M) and selected control proteins were measured using a competitive immunoassay to determine the origin of beta 2M in cows' milk. Using milk samples collected at various times, separated into different fractions and treated with protease inhibitors, it was established that beta 2M appears in cows' milk by protease-dependent degradation of the cellular fraction of milk, probably mononuclear cells, but is not derived from milk fat globules (MFG) or from polymorphonuclear leukocytes despite positive immunofluorescence of the former. The latter source could be eliminated by the induction of neutrophilia which produced no changes in beta 2M levels. Our data also indicate a minimal contribution by MFG to levels of secretory component despite its detection by indirect immunofluorescence on MFG but are consistent with the view that the milk fat globule membrane protein, bovine-associated microprotein, is derived from MFG by proteolysis. Protease-dependent degradation of milk components is an in vivo storage effect which occurs as early as the first 3 hr of in vivo storage. Compared to three other secretions tested, beta 2M was most concentrated in lacteal secretions and codistributed with bovine lymphocyte antigen in milk.

Animals↗
PubMed1985

In vivo proteolytic activity of the mammary gland. Contribution to the origin of secretory component, beta 2-microglobulin and bovine-associated mucoprotein (BAMP) in cows milk.

Milk samples were collected from Holstein-Friesian cows at various times after milking (10-30 min; 30 min-10 hr) and treated with a protease inhibitor or control solution. Samples were then fractionated into whole, skimmed and cell-free skimmed milk aliquots. Some animals were treated with E. coli endotoxin prior to sample collection. The concentrations of three membrane-associated proteins (MAP), beta 2-microglobulin (beta 2M), secretory component (SC) and bovine-associated mucoprotein (BAMP) as well as albumin were measured in each aliquot to determine if in vivo proteolysis of milk elements could explain the origin of these MAP in milk. All three MAP could be localized on milk fat globules (MFG) and alveolar epithelial cells of the gland. Data revealed that all BAMP in milk can be accounted for by in vivo proteolytic degradation of MFG while most beta 2M is derived by similar degradation, from cellular elements in milk, presumably monocytes. Experiments with endotoxin which elevate PMN levels, failed to influence the release of any MAP while elevating albumin levels by greater than 10-fold. Based on these studies, SC release into milk cannot be ascribed to a protease-dependent mechanism.

Animals↗
PubMed1984

Production and characterization of monoclonal bovine immunoglobulins G1, G2 and M from bovine x murine hybridomas.

Three bovine x murine hybridomas, which secrete bovine IgG1, IgG2 and IgM, respectively, were derived by the fusion of normal bovine B lymphocytes to the murine cell line SP-2/0. The hybridomas were initially selected by testing the culture supernatant of individual clones with rabbit anti-bovine light chain antibody. The bovine nature of Ig secreted by the bovine x murine hybridomas was confirmed by the ability of bovine serum but not murine serum to inhibit specific adsorption by murine anti-bovine Ig coupled to Sepharose 4B and the inability of sheep anti-mouse Ig (all isotypes) to bind biosynthetically labelled Ig from the respective bovine x murine hybridomas or to react with bovine x murine hybridoma Ig in Ouchterlony analysis. The isotype of the bovine Ig produced by each hybridoma was determined by cRIA using bovine Ig isotype-specific murine mcab, Ouchterlony analysis with guinea pig anti-bovine Ig isotypes and molecular weight analysis of unreduced and reduced affinity purified Ig from the respective bovine x murine hybridomas. Even though the hybridomas in this study showed chromosome loss, they have continued to secrete bovine Ig in culture for over 16 months, indicating that it is possible to isolate and stabilize interspecific hybridomas retaining the chromosome, or at least the genes, encoding an Ig. These hybridomas will provide monoclonal bovine Ig for; 1) serological standards, 2) production of polyclonal and monoclonal antisera to bovine Ig isotypes, 3) sequencing studies, 4) serological and structural studies of bovine Ig classes and subclasses, and messenger RNA for the production of cDNA probes for the cloning of bovine Ig genes and for determining the organization of Ig genes in the bovine genome.

Animals↗
PubMed1984

Characterization of monoclonal antibodies to bovine IgG1.

Monoclonal antibodies were developed to bovine IgG1. In addition, production of monoclonal antibodies to bovine light chain is also reported. Monoclonal antibody specificities were initially determined by solid-phase enzyme immunoassay. The monoclonal antibovine IgG1 was shown by a specificity-independent isotyping solid-phase enzyme immunoassay to be mouse IgG1 with kappa light chains. Ascites derived monoclonal antibovine IgG1 antibodies were linked to cyanogen bromide-activated Sepharose and used for affinity isolation of bovine IgG1. The bovine IgG1 eluted from the affinity column was characterized using immuno-electrophoresis, acrylamide gel electrophoresis, isoelectric focusing and solid-phase enzyme immunoassay. Affinity chromatography using monoclonal antibodies provided both a verification of monoclonal antibody specificity and a rapid technique for the isolation of bovine IgG1. This technique may also be employed to remove IgG1 contaminants during purification of bovine IgA.

Animals↗
PubMed1983

Bovine x mouse hybridomas that secrete bovine immunoglobulin G1.

The interspecific fusion of normal bovine lymphocytes with a nonsecreting mouse hybridoma produced stable cell lines secreting bovine immunoglobulins. One of these lines has continued to secrete immunoglobulin G1 (5 to 10 micrograms per milliliter) for over 16 months. The bovine x mouse hybrid cells can be expected to provide bovine monoclonal immunoglobulins for sequencing studies and for use as serological standards as well as to provide messenger RNA for cloning bovine immunoglobulin genes.

Animals↗
PubMed1983

Production of monoclonal antibodies to bovine leukocyte cell-surface components.

Fusion of murine myeloma cells with syngeneic spleen lymphocytes has led to development of hybridomas secreting antibodies. Hybrid cells retain the immortality and clonability of myeloma parents as well as the antibody-producing property of lymphocytes. Specificity of monoclonal antibody produced is based on the one lymphocyte-one antibody phenomenon and represents the most effective process for producing specific antisera. spleen cells from mice immunized with desired antigen are hybridized with nonsecreting mouse myeloma cells. Resulting hybrids are cloned and culture fluids tested for specific antibody activity to the antigen. Positive clones are cultivated in vitro and injected into mice for monoclonal antibody production. This technology has been extended to the bovine species to obtain monoclonal antisera to immunoglobulins and cell-surface components of leukocytes for study of mammary gland immunity. Recent progress in monoclonal research has led to interspecific fusion of murine myelomas with bovine lymphocytes, resulting in hybridomas that produce monoclonal bovine immunoglobulins. Monoclonal antibodies will be useful in investigations applicable to bovine research including purification of immunoglobulins, determining immunoglobulin concentration in colostrum and milk, reference reagents for bovine serology, antibody localization in tissue, gene sequencing, characterizing histocompatibility antigens, distinguishing and quantitating cell types in blood, milk, and udder tissue, and elucidating role of cell subpopulations in the immune response.

Animals↗
PubMed1983

Sequential response of milk leukocytes, albumin, immunoglobulins, monovalent ions, citrate, and lactose in cows given infusions of Escherichia coli endotoxin into the mammary gland.

Changes in concentrations of both the cellular and the humoral components of milk are known to occur during mastitis. This study was conducted to determine temporal changes in the concentrations of leukocytes, albumin, immunoglobulins (Ig), monovalent ions, lactose, and citrate in milk during the initial phases of simulated mastitis. Ten cows whose udders were pathogen free and had milk leukocyte counts of less than 0.5 X 10(6)/ml were used. Two dosages of Escherichia coli endotoxin were administered to simulate various degrees of mastitis. Two quarters in each cow were infused with the endotoxin and the other 2 served as controls. Quarter milk samples were collected frequently before and after infusion. Within 2 hours after infusion of a 100-micrograms dose of endotoxin, clinical mastitis was observed in most of the infused quarters. Leukocytes, albumin, IgG1, and conductivity showed significant increases. Values before infusion and at postinfusion (PI) hour 2 were as follows: leukocytes, 0.33 and 3.65 X 10(6)/ml, respectively; albumin, 0.38 and 4.49 mg/ml; IgG1, 0.34 and 0.79 mg/ml; and conductivity, 6.0 and 6.9 mmho. Average of the peak values and their average relative time of appearance after infusion were as follows: leukocytes, 28.82 X 10(6)/ml at 16 hours; albumin, 9.37 mg/ml at 4 hours; IgG1, 1.35 mg/ml at 4 hours; and conductivity, 95.5 mmho at 10 hours. The IgG1 values tended to remain high in the presence of rapidly declining albumin concentrations, indicating the possibility of an active, rather than a passive, transfer of IgG1 from the circulation. The response to the 10-micrograms dose of endotoxin ranged from subclinical to clinically mild mastitis with lesser cellular and humoral responses.

Animals↗
PubMed1982

Production and characterization of monoclonal antibodies to bovine immunoglobulin G2.

Spleen cells from Balb/cJ mice inoculated with bovine immunoglobulin (Ig)G were hybridized with hypoxanthine aminopterin thymidine-sensitive nonsecreting cell line SP-2/0, and the hybrid cell culture fluid was tested for specificity. Hybrid cells secreting monoclonal antibodies to bovine IgG were isolated and recloned. The monoclonal antibody DAS 2 was specific for bovine IgG2 by an indirect solid-phase radioimmunoassay. This clone was adapted for growth as antibody-secreting neoplasms in Balb/cJ mice, the sera from which contained high titers (1:128,000) of anti-bovine IgG2 antibodies. Antibody specific to bovine IgG2 was isolated by affinity chromatography. This antibody was shown by Ouchterlony analysis to be mouse IgG1 with the kappa light chain. On isoelectric focusing, this antibody gave a pattern that was consistent with monoclonality.

Animals↗
PubMed1981

Concentrations of corticosteroids, leukocytes, and immunoglobulins in blood and milk after administration of ACTH to lactating dairy cattle: effects on phagocytosis of Staphylococcus aureus by polymorphonuclear leukocytes.

A study was conducted to determine relationships among plasma and milk corticosteroids, milk immunoglobulins, and phagocytosis of Staphylococcus aureus by polymorphonuclear leukocytes (PMN) isolated from milk of cows given injections of 0 (saline solution only), 100, and 200 IU of ACTH. Also determined were the effects of ACTH on mobilization of PMN into milk from the mammary gland irritated by infusion of 100 ml of saline solution containing 0.1% oyster glycogen. Cows (n = 10) were injected 6 times within a 48-hour period with 0, 100, or 200 IU of ACTH. Immediately before cows were given the 5th injection, 2 mammary quarters were infused with the saline-glycogen solution. Five hours after the 6th injection, milk from infused quarters was collected, and PMN were isolated, washed, and resuspended in autologous skimmed milk collected 5 hours after the 4th injection and before the udder was infused. Isolated PMN were incubated with S aureus and percentage of phagocytosis was determined. Concentrations of total corticosteroids in plasma and milk increased after cows were given injections of 100 and 200 IU of ACTH. The concentrations of IgA, IgG1, IgG2, IgM, and bovine serum albumin in milk after 4 injections of 0, 100, and 200 IU of ACTH were similar to preinjection (base line) concentrations. Injections of 100 and 200 IU of ACTH significantly increased the concentrations of total circulating leukocytes. Concentrations of leukocytes in milk tended to increased with increasing doses of ACTH, but the differences were not significant. Injection of 100 and 200 IU of ACTH reduced phagocytosis of S aureus by PMN. After 60 minutes of incubation, phagocytosis averaged 57% and 54%, respectively, for the ACTH treatments and 70% for controls (saline only). Results indicate that injections of ACTH that result in physiologic and pharmacologic plasma concentrations of corticosteroids inhibit phagocytosis. Impairment of phagocytosis appeared to be a direct effect of corticosteroid concentration o PMN and was not due to reduced concentrations of immunolobulins. These data indicate that reduced phagocytosis by PMN could be important in increased susceptibility to disease during stress in lactating dairy cows.

Adrenal Cortex Hormones↗
PubMed1980

Quarter milk variation in immunoglobulins and ability to support phagocytosis.

Milk samples were collected from two quarters in each of 8 cows that had Wisconsin Mastitis Test scores of less than 8.0. After letdown was stimulated, 35 ml of foremilk, the sixth 35 ml (middle milk), and 45 ml of strippings were collected by hand. Measured on each fraction were immunoglobulins, serum albumin, total protein, pH, percentage fat, Wisconsin Mastitis Test, and the ability of skim milk and whole milk to support phagocytosis of Staphylococcus aureus by isolated donor milk neutrophils. Foremilk and middle milk differed in immunoglobulin M (.014 to .018 mg/ml) and in the ability of whole milk to support phagocytosis (32 to 28% S. aureus phagocytosed). The following measures differed from middle milk to strippings: immunoglobulin A (.063 to .083 mg/ml); immunoglobulin G1 (.238 to .302 mg/ml); immunoglobulin M (.018 to .023 mg/ml); fat (3.9 to 7.4%); and Wisconsin Mastitis Test (5.0 to 9.2). Immunoglobulin G2, bovine serum albumin, total protein, pH, and the ability of skim milk and whole milk to support phagocytosis did not change. Although small differences may exist between foremilk and middle milk, foremilk gave reliable estimates.

Animals↗
PubMed1980

Effect of local immunization of the mammary gland on phagocytosis and intracellular kill of Staphylococcus aureus by polymorphonuclear neutrophils.

Four cows in the latter part of their 2nd, 3rd, or 4th lactations were immunized by multiple intramammary infusions of heat-killed Staphylococcus aureus in 2 quarters. The direct bactericidal effects of milk whey from the immunized and control quarters, before and after immunization, and the ability of these whey to support phagocytosis and intracellular kill were determined by incubating live S aureus with polymorphonuclear neutrophils isolated from milk. Immunoglobulins (Ig) were determined by single radial immunodiffusion. One immunized and 1 control quarter in each cow were challenge exposed with live S aureus and the courses of the infections were determined for 2 weeks. There were significant cow differences in all Ig classes and in percentage of phagocytosis. Immunization resulted in a significant increase in IgA, IgG2, and IgM in the immunized quarters. Whey collected from immunized quarters supported phagocytosis of S aureus by isolated milk polymorphonuclear neutrophils significantly greater than did whey from control quarters. Extracellular live S aureus in the incubation medium was decreased by 59% in whey collected after immunization from immunized quarters. This decrease in extracellular S aureus was associated with a concomitant increase in total intracellular S aureus. However, intracellular live organisms showed no change. This lack of change indicated that the additional S aureus that were phagocytosed were killed. Direct bactericidal effects of whey were not observed. Intracellular live S aureus was not significantly correlated with any of the variables measured.

Agglutination Tests↗
PubMed1980

Effect of udder inflammation on milk immunoglobulins and phagocytosis.

Udder inflammation was induced in two quarters in each of eight cows by infusing oyster glycogen in saline solution. Milk was collected before and after infusion for immunoglobulin (Ig) and phagocytosis assays. Milk Ig and bovine serum albumin were measured by single radial immunodiffusion. The concentrations (mg/ml) of the immunoglobulins (IgA, IgG1, IgG2, and IgM) and bovine serum albumin before and after infusion were: IgA = 0.064 (before), 0.084 (after); IgG1 = 0.323, 0.486; IgG2 = 0.046, 0.094; IgM = 0.091, 0.124; and bovine serum albumin = 0.207, 0.451. The fact that IgA, IgG1, IgG2, and IgM were significantly correlated with bovine serum albumin within the cows before and after inflammation suggests a passive transfer of these Ig during inflammation. Phagocytosis was determined by incubating polymorphonuclear leukcotyes collected after infusion with 32P-tagged Staphylococcus aureus in autologous skim milk collected before and after infusion. The difference in percentage of phagocytosis before (42%) and after (36%) infusion was not significant. None of the Ig were significantly correlated with phagocytosis.

Animals↗
PubMed1980

Relationship among leukocyte phagocytosis, milk immunoglobulins, and susceptibility to intramammary infection.

A study was conducted to determine the variation among cows (n = 7 and among quarters within cows in the ability of milk to support phagocytosis, immunoglobulin (Ig) concentration, and the response to intramammary challenge exposure with live Staphylococcus aureus. Before challenge exposure, the average concentration (milligrams/milliliter) of each class of Ig was: IgA = 0.05, IgG1 = 0.42, IgG2 = 0.06, and IgM = 0.05. The percentage of phagocytosis of S aureus by milk neutrophils was significantly correlated with IgA (r = 0.59; P < 0.01), IgG1 (r = 0.52; P < 0.01), and agglutination titer (r = 0.68; P < 0.01). Agglutination titer was significantly correlated with IgA (r = 0.52; P < 0.01) and IgG1 (r = 0.63; P < 0.01). Bovine serum albumin was significantly correlated with IgA (r = 0.55; P < 0.01) and IgG2 (r = 0.49; P < 0.01). An IgG1/IgG2 ratio of 7.0 in milk, as compared with an IgG1/IgG2 ratio of 0.7 in serum, indicated a selective transport for IgG1. After challenge exposure, the IgG1/IgG2 ratio in infected and noninfected quarters was 12.9 and 16.0, respectively. The fact that neither the bovine serum albumin nor the direct microscopic somatic cell count in the noninfected quarters changed indicated that the increase in the IgG1/IgG2 ratio was due to an increase in selective transport in all quarters and not to a change in permeability due to a generalized inflammatory response. The correlation between the number of challenge exposures required to produce an infection was significantly correlated with percentage of phagocytosis (r = 0.70; P < 0.01).

Agglutination Tests↗
PubMed1979

Leukocytes--second line of defense against invading mastitis pathogens.

In mammals, neutrophile polymorphonuclear leukocytes constitute one of the essential body defenses against disease. In a large mammal, such as the dairy cow, billions of neutrophils are mobilized to fight infection. For example, over 50 million neutrophils per milliliter milk are commonly in a mammary quarter inflicted with clinical mastitis. However, in spite of these numerous leukocytes, pathogenic organisms remain viable. Recent evidence indicates that bacteria are not eliminated from a diseased quarter because the phagocytic capacity of the neutrophils is reduced in the mammary gland. The morphology and physiology of the leukocyte is examined in this review in an attempt to explain why the phagocytic capacity of the neutrophil is reduced in the mammary gland of the bovine.

Animals↗