New carbohydrate-active enzymes identified by screening two metagenomic libraries derived from the soil of a winter wheat field.

AIMS: Soils are rich, diversified environments where ß-glucosidases abound because of their importance in organic matter degradation. The aim of this work was to discover new ß-glucosidases by constructing two metagenomic DNA libraries from soil samples collected in winter and spring from a field of winter wheat. METHODS AND RESULTS: Both libraries were screened on esculin-supplemented medium so as to isolate candidates showing ß-glucosidase activity. Candidate analysis revealed seven putative ß-glycosidases and two putative glycosyltransferases, displaying 25 to 82% identity to known enzymes. The putative ß-glycosidases belong to families GH1, GH3 and GH20 and the two putative glycosyltransferases, probably, to new families. In characterization tests performed on bacteria in suspension or spread on agar plates, some candidates appeared to hydrolyse several natural and synthetic substrates. These tests also highlighted interesting industrial characteristics, such as the activity of four ß-glycosidases under alkaline conditions and the esculin-hydrolysing activity of a ß-glucosidase candidate in the presence of glucose. CONCLUSIONS: Seven putative ß-glycosidases and two putative glycosyltransferases were found by functional screening of two metagenomic DNA libraries derived from agricultural soil. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has identified ß-glycosidases and putative glycosyltransferases that have or may have interesting industrial characteristics.

Bovine leukemia virus-related antigens in lymphocyte cultures infected with AIDS-associated viruses.

An earlier finding that lymphocytes from African patients with the acquired immune deficiency syndrome (AIDS) react with rabbit antiserum to purified antigens of bovine leukemia virus (BLV) prompted a study of the possible cross-reactions between a BLV-infected ovine cell line and human lymphocytes inoculated with a strain of lymphadenopathy syndrome-associated virus (LAV). A solid-phase radioimmunoassay was used to detect antigenic markers of the retroviruses. Crude extracts from short-term cultures of lymphocytes infected with LAV bound rabbit antisera to the LAV glycoprotein gp13 (molecular weight 13,000) and the BLV proteins p24 and gp51, but did not bind antibodies to the p24 of human T-cell leukemia virus type I (HTLV-I). Antiserum to LAV gp13 reacted with an ovine cell line producing BLV but also weakly with virus-free ovine cells. Lymphocyte cultures from four African patients with AIDS expressed BLV-related antigens within 6 to 10 days of culture, at the moment when particle-bound reverse transcriptase was produced. BLV-related antigens were induced in lymphocyte cultures from healthy individuals by addition of filtered supernatant or irradiated cells of the original culture. The antisera to BLV used in this study may prove useful for the detection of AIDS-associated viruses in short-term cultures of lymphocytes from AIDS patients or their contacts.

Replicative and cytopathic potential of HTLV-III/LAV with sor gene deletions.

The genome of the human T-lymphotropic virus type III (HTLV-III/LAV) has the potential to encode at least three polypeptides in addition to those encoded by the gag, pol, and env genes. In this study, the product of the sor (short open reading frame) region, which overlaps the 3' end of the pol gene, was found to be a protein with a molecular weight of 23,000. An assay was developed for testing the ability of cloned HTLV-III proviruses to produce viruses cytopathic for T4+ lymphocytes. In the cell line used, C8166, neither the HTLV-III sor gene product nor the complete 3'-orf gene product were necessary for the replication or cytopathic effects of the HTLV-III.

The composite genome of the legume symbiont Sinorhizobium meliloti.

The scarcity of usable nitrogen frequently limits plant growth. A tight metabolic association with rhizobial bacteria allows legumes to obtain nitrogen compounds by bacterial reduction of dinitrogen (N2) to ammonium (NH4+). We present here the annotated DNA sequence of the alpha-proteobacterium Sinorhizobium meliloti, the symbiont of alfalfa. The tripartite 6.7-megabase (Mb) genome comprises a 3.65-Mb chromosome, and 1.35-Mb pSymA and 1.68-Mb pSymB megaplasmids. Genome sequence analysis indicates that all three elements contribute, in varying degrees, to symbiosis and reveals how this genome may have emerged during evolution. The genome sequence will be useful in understanding the dynamics of interkingdom associations and of life in soil environments.

Genomic location of the bovine growth hormone secretagogue receptor (GHSR) gene and investigation of genetic polymorphism.

The growth hormone secretagogue receptor (GHSR) is involved in the regulation of energetic homeostasis and GH secretion. In this study, the bovine GHSR gene was mapped to BTA1 between BL26 and BMS4004. Two different bovine GHSR CDS (GHSR1a and GHSR1b) were sequenced. Six polymorphisms (five SNPs and one 3-bp indel) were also identified, three of them leading to amino acid variations L24V, D194N, and Del R242. These variations are located in the extracellular N-terminal end, the exoloop 2, and the cytoloop 3 of the receptor, respectively.

Genetic analysis of lactoferrin content in bovine milk.

Bovine lactoferrin (LF) is mainly present in milk and shows important physiological and biological functions. The aim of this study was to estimate the heritability and correlation values of LF content in bovine milk with different economic traits as milk yield (MY), fat and protein percentages, and somatic cell score (SCS). Variance components of the studied traits were estimated by REML using a multiple-trait mixed model. The obtained heritability (0.22) for LF content predicted using mid-infrared spectrometry (pLF) suggested the possibility of animal selection based on the increase of LF content in milk. The phenotypic and genetic correlation values calculated between pLF and SCS were moderate (0.31 and 0.24, respectively). Furthermore, a preliminary study of bovine LF gene polymorphism effects was performed on the same production traits. By PCR, all exons of the LF gene were amplified and then sequenced. Three new polymorphisms were detected in exon 2, exon 11, and intron 8. We examined the effects of LF gene polymorphisms of exons 2, 4, 9, 11, and 15, and intron 8 on pLF, MY, fat and protein percentages, and SCS. The different observed effects did not reach a significant level probably because of the characteristics of the studied population. However, the results were promising, and LF may be a potential indicator of mastitis. Further studies are necessary to evaluate the effect of genetic selection based on LF content on the improvement of mastitis resistance.

Effects of feed supplementation with specific hen egg yolk antibody (immunoglobin Y) on Salmonella species cecal colonization and growth performances of challenged broiler chickens.

Anti-Salmonella spp. egg yolk antibodies (IgY) simultaneously directed against Salmonella Enteritidis and Salmonella Typhimurium were tested to determine if their inclusion in feed decreased Salmonella spp. cecal colonization in experimentally infected broiler chickens. Egg yolk powder (EYP) was obtained by freeze-drying egg yolks containing anti-Salmonella spp. Immunoglobin Y was included in feed at 5 levels of concentration (0 to 5%). Feeds were formulated to similar nutrient levels and provided for ad libitum intake from d 1 to 28. Three days after initiation of feed treatments (d 4), chickens were co-challenged with equal numbers of Salmonella Enteritidis and Salmonella Typhimurium (2x10(6) cfu/bird). Cecal samples were recovered weekly over the experimental period (d 7 to 28) to enumerate Salmonella spp. The effect of anti-Salmonella spp. IgY feed supplementation on growth performance of infected chickens was also evaluated during the same period. In comparison with the positive control treatment (PC), treatments involving EYP (T1, T2, T3, T4, and T5), whether containing anti-Salmonella spp. IgY or not, significantly improved (P<0.05) the growth performance of challenged chickens, but without reaching the performance levels of nonchallenged chickens (NC1 and NC2). However, no link can be established between the enhancement in growth performance of challenged birds and their contamination levels by Salmonella because in-feed incorporation of EYP had no significant effect on cecal colonization by Salmonella. Furthermore, the comparison of the 5 anti-Salmonella spp. IgY concentration levels in feed did not reveal any anti-Salmonella spp. IgY concentration effect on growth performance and Salmonella cecal colonization. These results suggest that anti-Salmonella spp. IgY would undergo denaturation and degradation after their passage through the animal gastrointestinal tract and reveal that components of EYP other than specific antibodies have a beneficial effect on growth performance.

Production of hen egg yolk immunoglobulins simultaneously directed against Salmonella enteritidis and Salmonella typhimurium in the same egg yolk.

The present study was an attempt to raise hen egg yolk Ig (IgY) simultaneously directed against Salmonella Enteritidis (SE) and Salmonella Typhimurium (ST) in the same egg yolk. The immunopotentiating effect of 2 different adjuvants -- Freund's adjuvants (FA) and immunostimulating complexes matrix (IM) -- on antibody response was also evaluated. Bacterial outer membrane proteins (OMP) were selected as target antigens. The ISA Brown hens, specific-Salmonella spp.-free status, divided into 6 groups were intramuscularly injected with a mono-compound antigen preparation: SE-OMP (treatment SE-FA or SE-IM) or ST-OMP (treatment ST-FA or ST-IM), or a combined antigen preparation: (1/2) SE-OMP and (1/2) STOMP (treatment SEST-FA or SEST-IM). Titers of antibodies in yolk were evaluated biweekly with ELISA. There was no antigen x adjuvant interaction on antibody titers. Anti-SE IgY titers in hens that received treatment SEST-FA or SEST-IM were statistically similar (P > 0.05) as compared with those obtained from hens immunized with treatment SE-FA or SE-IM. Anti-ST IgY titers in hens immunized with treatment SEST-FA or SEST-IM were slightly lower than those of hens that received treatment ST-FA or ST-IM. The cross-reactivity of anti-SE IgY, induced by treatment SE-FA or SE-IM, with ST-OMP antigen and that of anti-ST IgY, induced by ST-FA or ST-IM, with SE-OMP antigen were arbitrarily assessed on d 43 and 155 by ELISA. The average cross-reactivity of anti-SE IgY with ST-OMP antigen was 71.7%. The average cross-reactivity of anti-ST IgY with SE-OMP antigen was 78.8%. In FA groups, antibody titers were found higher (P < 0.05) than those in IM groups. Furthermore, no extensive lesions or clinical abnormalities were detected in hens injected with FA. These findings showed the opportunity to raise IgY antibody against 2 Salmonella serovars in the same yolk and that FA was more efficient than IM in mediating antibody response.

Reducing agent can be omitted in the incubation medium of the batch in vitro fermentation model of the pig intestines.

Over the past decade, in vitro methods have been developed to study intestinal fermentation in pigs and its influence on the digestive physiology and health. In these methods, ingredients are fermented by a bacterial inoculum diluted in a mineral buffer solution. Generally, a reducing agent such as Na2S or cysteine-HCl generates the required anaerobic environment by releasing metabolites similar to those produced when protein is fermented, possibly inducing a dysbiosis. An experiment was conducted to study the impact of two reducing agents on results yielded by such in vitro fermentation models. Protein (soybean proteins, casein) and carbohydrate (potato starch, cellulose) ingredients were fermented in vitro by bacteria isolated from fresh feces obtained from three sows in three carbonate-based incubation media differing in reducing agent: (i) Na2S, (ii) cysteine-HCl and (iii) control with a mere saturation with CO2 and devoid of reducing agent. The gas production during fermentation was recorded over 72 h. Short-chain fatty acids (SCFA) production after 24 and 72 h and microbial composition of the fermentation broth after 24 h were compared between ingredients and between reducing agents. The fermentation residues after 24 h were also evaluated in terms of cytotoxicity using Caco-2 cell monolayers. Results showed that the effect of the ingredient induced higher differences than the reducing agent. Among the latter, cysteine-HCl induced the strongest differences compared with the control, whereas Na2S was similar to the control for most parameters. For all ingredients, final gas produced per g of substrate was similar (P>0.10) for the three reducing agents whereas the maximum rate of gas production (R max) was reduced (P0.10) after 24 h of fermentation with Na2S and in the control without reducing agent. Molar ratios of branched chain-fatty acids were higher (P<0.05) for protein (36.5% and 9.7% for casein and soybean proteins, respectively) than for carbohydrate (<4%) ingredients. Only fermentation residues of casein showed a possible cytotoxic effect regardless of the reducing agent (P<0.05). Concerning the microbial composition of the fermentation broth, most significant differences in phyla and in genera ascribable to the reducing agent were found with potato starch and casein. In conclusion, saturating the incubation media with CO2 seems sufficient to generate a suitable anaerobic environment for intestinal microbes and the use of a reducing agent can be omitted.

Genetic variability of lactoferrin content estimated by mid-infrared spectrometry in bovine milk.

The effects of lactoferrin (LF) on the immune system have already been shown by many studies. Unfortunately, the current methods used to measure LF levels in milk do not permit the study of the genetic variability of lactoferrin or the performance of routine genetic evaluations. The first aim of this research was to derive a calibration equation permitting the prediction of LF in milk by mid-infrared spectrometry (MIR). The calibration with partial least squares on 69 samples showed a ratio of standard error of cross-validation to standard deviation equal to 1.98. Based on this value, the calibration equation was used to establish an LF indicator trait (predicted LF; pLF) on a large number of milk samples (n = 7,690). A subsequent study of its variability was conducted, which confirmed that stage of lactation and lactation number influence the overall pLF level. Small differences in mean pLF among 7 dairy breeds were also observed. The pLF content of Jersey milk was significantly higher than that in Holstein milk. Therefore, the choice of breed could change the expected LF level. Heritability estimated for pLF was 19.7%. The genetic and phenotypic correlations between somatic cell score and pLF were 0.04 and 0.26, respectively. As somatic cell score increases in presence of mastitis, this observation seems to indicate that pLF, or a function of observed pLF, compared with expected LF might have potential as an indicator of mastitis. The negative genetic correlation (-0.36) between milk yield and pLF could indicate an undesirable effect of selection for high milk production on the overall LF level.

Effects of oral supplementation with bovine colostrum on the immune system of weaned piglets.

The aim of this study was to evaluate the influence of bovine colostrum supplementation on the immune system of weaned piglets in a context of a full ban of in-feed antibiotics. After weaning at 21 days, 24 outbred piglets were fed with a diet supplemented daily for three weeks with 0, 1 or 5 g of colostrum. Feed intake, growth performance, haematological parameters, and serum and local anti-colostrum immunoglobulin levels were examined. Lymphocytes from the blood, spleen, and gut-associated lymphoid were analysed for phenotype as well as for their ability to produce cytokines. The stimulation index (SI) of mononuclear cells from different organs was obtained after colostral or mitogenic stimulation. Feed intake, growth, and haematological parameters were not significantly affected by colostrum. Total serum IgA levels were increased after colostrum supplementation, with a transient decrease in total IgG. Local anti-colostrum immunization was observed in colostrum-fed piglets. The CD21+/CD3+ cells populations of the ileal Peyer's patch (iPP) were markedly affected. The SI of lymphocyte populations changed significantly whereas, naive blood lymphocytes were not stimulated in vitro in the presence of bovine colostrum, suggesting local anti-colostrum immunization and an absence of direct mitogenic effects of the colostrum. Both Th1 and Th2 cytokine production was present in the different organs of colostrum-fed piglets. Bovine colostrum especially stimulated iPP cells.

Humoral antibody response to bovine leukemia virus infection in cattle and sheep.

In this study, 345 cattle from 7 herds with a history of lymphosarcoma were tested for antibody to BLV antigens by three serological methods, namely immunodiffusion using a bovine leukemia virus glycoprotein with a molecular weight of 60,000 as antigen, and radioimmunoassay using a bovine leukemia virus glycoprotein with a molecular weight of 60,000 and a bovine leukemia virus protein with a molecular weight of 24,000 as antigen. The three tests under comparison agreed for 335 animals, 240 being negative in the three tests, and 95 being positive. Results were variable in ten cases only. Glycoprotein with a molecular weight of 60,000 antibody titers were systematically higher than were protein with a molecular weight of 24,000 antibody titers in bovine sera and milk, as well as in sera of experimentally infected sheep. In the latter case, antibodies to bovine leukemia virus antigens reached maximal values at the animal death in the tumor phase of the disease. Ratios of serum antiglycoprotein titer to milk titer varied between 4 and 117, showing that, if milk pools are to be used in surveys of bovine leukemia virus infection, use of very sensitive techniques of detection is mandatory.

Adding mucins to an in vitro batch fermentation model of the large intestine induces changes in microbial population isolated from porcine feces depending on the substrate.

Adding mucus to in vitro fermentation models of the large intestine shows that some genera, namely lactobacilli, are dependent on host-microbiota interactions and that they rely on mucosal layers to increase their activity. This study investigated whether this dependence on mucus is substrate dependent and to what extent other genera are impacted by the presence of mucus. Inulin and cellulose were fermented in vitro by a fecal inoculum from pig in the presence or not of mucin beads in order to compare fermentation patterns and bacterial communities. Mucins increased final gas production with inulin and shifted short-chain fatty acid molar ratios (P < 0.001). Quantitative real-time PCR analyses revealed that Lactobacillus spp. and Bifidobacterium spp. decreased with mucins, but Bacteroides spp. increased when inulin was fermented. A more in-depth community analysis indicated that the mucins increased Proteobacteria (0.55 vs 0.25%, P = 0.013), Verrucomicrobia (5.25 vs 0.03%, P = 0.032), Ruminococcaceae, Bacteroidaceae and Akkermansia spp. Proteobacteria (5.67 vs 0.55%, P < 0.001) and Lachnospiraceae (33 vs 10.4%) were promoted in the mucus compared with the broth, while Ruminococcaceae decreased. The introduction of mucins affected many microbial genera and fermentation patterns, but from PCA results, the impact of mucus was independent of the fermentation substrate.

Cloning and characterization of MN, a human tumor-associated protein with a domain homologous to carbonic anhydrase and a putative helix-loop-helix DNA binding segment.

MN is a transmembrane glycoprotein that has been detected in HeLa cells and in some human carcinomas. The expression of MN protein in HeLa cells is regulated by cell density. In HeLa x fibroblast cell hybrids its expression correlates with tumorigenicity. Using a specific monoclonal antibody we have identified a cDNA clone coding for MN. Analysis of the deduced amino acid sequence revealed strong structural homology between the central region of the MN protein and carbonic anhydrases (CA). MN sequence retains the conserved zinc-binding site as well as the enzyme's active center. In accord with these findings, MN protein from HeLa cells was found to bind zinc and to have carbonic anhydrase activity. The N-terminal region of MN shares some similarity with DNA binding proteins of the helix-loop-helix (HLH) family, and the protein was found to have affinity for DNA by DNA-cellulose chromatography. The region between the CA-like domain and the putative HLH domain is rich in imperfect repeats of serine, proline, glycine and acidic residues with few hydrophobic amino acids, resembling thus an activation region of transcription factors. The fact that MN protein is detectable in several types of human carcinomas, but not in corresponding non-cancerous tissues, suggests its possible role in neoplasia. In addition, the analysis of biological consequences of MN expression of NIH3T3 cells provides the evidence in favour of MN protein involvement in control of cell proliferation and transformation.

Phosphorylation of bovine leukemia virus Tax protein is required for in vitro transformation but not for transactivation.

The Tax proteins of the oncovirinae viruses are phosphorylated transcriptional activators that exhibit oncogenic potential. The role of phosphorylation in their functional activities remains unknown. As a model for the Human T-cell leukemia virus type I (HTLV-I), Bovine Leukemia Virus (BLV) permits the characterization of viral replication and leukemogenesis in vivo. Here, we show that the BLV Tax protein is phosphorylated on serine residues 106 and 293 both in insect and in mammalian cells. These sites can also be efficiently phosphorylated by the cdc2 and MAP kinases in vitro. Mutation of these residues does not affect the capacity of the Tax protein to function as a transactivator. Indeed, the Tax proteins mutated at one or both serines increase LTR-directed viral transcription at levels similar to those obtained with wild-type Tax in cell culture. Moreover, inhibition of Tax phosphorylation by W7, a calmodulin antagonist, does not alter its transactivation activity. Thus, phosphorylation on serines 106 and 293 is not required for transactivation by Tax. However, simultaneous substitution of both serines into alanine residues destroys the capacity of Tax to cooperate with the Ha-ras oncogene to transform primary rat embryo fibroblasts and induce tumors in nude mice. When the serines were replaced with aspartic acid residues, the oncogenic potential of Tax was maintained indicating that the negative charge rather than the phosphate group itself was required for Tax oncogenicity. Finally, to assess the role of the serine residues in vivo, recombinant viruses which express the Tax mutants were constructed and injected into sheep. It appeared that the mutated proviruses replicate at levels similar to the wild-type virus in vivo. We conclude that Tax phosphorylation is dispensable for transactivation and viral replication in vivo but is required for its oncogenic potential in vitro.

Experimental transmission of enzootic bovine leukosis to sheep: latency period of the tumoral disease.

In the field of viral oncogenesis the latency period is the interval between detectable establishment of infection and appearance of a tumor. Between 1969 and 1985, a total of 60 sheep died with lymphosarcoma. They were inoculated with BLV-positive blood from various donor cows, by various routes, at various ages, etc. A statistical analysis was performed trying to find a correlation between the length of the latency period and, on the other hand, one or more factors, such as sex, family lineage, identity of the dam, age at inoculation, route of inoculation, or origin of the inoculum. None of the above mentioned parameters has a significant effect on the length of the latency period. In two series of sheep inoculated with decreasing number of lymphocytes from BLV-positive donor cows, hematological disorders and tumors appeared at first in recipient animals inoculated with the higher doses of infectious blood. Thus, the inoculated dose has an effect upon the length of the latency period; the higher the dose inoculated, the shorter the latency period. This finding suggests an explanation to the natural occurrence of multiple case herds as opposed to no-tumor case herds. A multiple case herd fulfills two conditions: the presence of a good donor and an efficient route of transmission allowing the transfer to the recipient of the optimal amount of infected blood.

The bovine leukemia virus (BLV) envelope glycoprotein gp51 as a general model for the design of a subunit vaccine against retroviral infection: mapping of functional sites through immunological and structural data.

Further advances in retroviral vaccine development require a better understanding of the antigenic structure of the envelope complex which is directly involved in infectivity events such as receptor recognition and membrane fusion. To design an optimal vaccine against BLV infection, we chose an approach based on the use of synthetic peptides covering 78% of the gp51 sequence in order to select only those segments that could induce a protective response via cellular and humoral immunity. On the other hand, we built a model of the BLV env glycoprotein 3D organization, based upon the very sensitive hydrophobic cluster analysis (HCA). The major information highlighted from this model is that the two loops, against which the most efficient neutralizing antipeptides antibodies are directed against, are in close proximity at the top of the "head" and could represent a potential site for receptor binding. These two peptides are of particular interest since they induce also a helper T-cell response. We further propose that the BLV envelope glycoprotein oligomerizes as a trimer.

Biochemical and immunological characterization of the bovine leukemia virus (BLV) envelope glycoprotein (gp51) produced in Saccharomyces cerevisiae.

The nucleotide sequence coding for bovine leukemia virus (BLV) envelope glycoprotein gp51 was inserted into a yeast-Escherichia coli shuttle vector carrying the promoter and secretion signal sequence of PHO5 (the yeast gene coding for repressible acid phosphatase) and the CYC1 transcriptional terminator. Yeast cells transformed by this construction synthesized gp51 after PHO5 induction by inorganic phosphate deprivation. The yeast-expressed gp51 was partially glycosylated into heterodisperse protein molecules ranging from 40 to 48 kDa. No gp51 was excreted in the culture medium. The amount of protein accumulated in yeast cells was estimated to reach 0.06% of soluble proteins. This modest level of expression seemed to be due to the toxicity of gp51 to the yeast cell. The yeast-expressed gp51 products were used in enzyme-linked immunosorbent assays for the detection of antibodies in sera from BLV-infected animals; they were also screened for the presence of well-defined biological epitopes. In both studies poor reactivity was observed. Rabbits immunized with the recombinant gp51 showed high antibody titers to native BLV gp51. However, these antibodies did not neutralize BLV in vitro.

High yield synthesis of the bovine leukemia virus (BLV) p24 major internal protein in Saccharomyces cerevisiae.

Bovine leukemia virus (BLV) p24 gene was expressed in Saccharomyces cerevisiae under the control of the PHO5 (encoding repressible acid phosphatase, rAPase) promoter. Yeast cells were transformed by a yeast-E. coli shuttle vector carrying the PHO5 promoter, the p24 gene and the CYC1 transcription terminator. After low inorganic phosphate (Pi) induction of the PHO5 promoter, p24 accumulated in the producing cells up to a concentration representing 10% of total soluble proteins. The expression level of p24 gene was not increased by insertion of the positive regulatory gene PHO4 on the p24 expression vector. The p24 produced in this system and incubated in crude yeast extract showed a remarkably high resistance to proteolytic degradation, a feature that presumably correlates with the compact globular conformation of the protein combined to the stabilizing effect of the N-terminal residue.

Iodoacetamide treatment of bovine leukemia virus glycoprotein gp51 enhances the western blotting reactivity of anti-peptide antibodies.

Rabbit polyclonal antibodies were raised against synthetic peptides of the bovine leukemia virus envelope glycoprotein gp51 and tested against the full size protein by the Western blotting technique. We show that acetylation of gp51 by iodoacetamide either maintains or significantly increases the antigen-antibody reaction and conclude therefrom that the reactive potential of an anti-peptide antibody may require acetylation of the sulfhydryl groups of the blotted protein.