Commercial Fetal Bovine Serum Research Articles

Improving quality of analysis by suppression of unwanted signals through band-selective excitation in NMR spectroscopy for metabolomics studies.

Nuclear Magnetic Resonance (NMR) spectroscopy stands as a preeminent analytical tool in the field of metabolomics. Nevertheless, when it comes to identifying metabolites present in scant amounts within various types of complex mixtures such as plants, honey, milk, and biological fluids and tissues, NMR-based metabolomics presents a formidable challenge. This predicament arises primarily from the fact that the signals emanating from metabolites existing in low concentrations tend to be overshadowed by the signals of highly concentrated metabolites within NMR spectra. The aim of this study is to tackle the issue of intense sugar signals overshadowing the desired metabolite signals, an optimal pulse sequence with band-selective excitation has been proposed for the suppression of sugar's moiety signals (SSMS). This sequence serves the crucial purpose of suppressing unwanted signals, with a particular emphasis on mitigating the interference caused by sugar moieties' signals. We have implemented this comprehensive approach to various NMR techniques, including 1D 1H presaturation (presat), 2D J-resolved (RES), 2D 1H-1H Total Correlation Spectroscopy (TOCSY), and 2D 1H-13C Heteronuclear Single Quantum Coherence (HSQC) for the samples of dates-flesh, honey, a standard stock solution of glucose, and nine amino acids, and commercial fetal bovine serum (FBS). The outcomes of this approach were significant. The suppression of the high-intensity sugar signals has considerably enhanced the visibility and sensitivity of the signals emanating from the desired metabolites. This, in turn, enables the identification of a greater number of metabolites. Additionally, it streamlines the experimental process, reducing the time required for the comparative quantification of metabolites in statistical studies in the field of metabolomics.

Analysis of commercial fetal bovine serum (FBS) and its substitutes in the development of cultured meat

Fetal bovine serum (FBS) is an extremely important culture growth supplement, accounting for approximately 60 % of cell-culture-media costs; therefore, lowering FBS-acquisition costs for the industrialization of cultured meat is imperative. This study attempted to produce an FBS substitute using discarded livestock by-products, with particular focus on formulating a product with a composition similar to that of FBS to improve effectiveness. However, to date, no study has precisely analyzed the commercial components of FBS, and this study is the first to compare the chemical composition of FBS and commercially available horse serum purchased from the United States or Europe with that of FBS substitutes developed by our team. This study analyzed the chemical composition of the FBS products purchased by our team over the past 3 years via blood, sodium dodecyl sulfate–polyacrylamide gel electrophoresis, and independent composition analyses. While the composition and quality of commercial FBS products are known to vary, the FBS composition of our purchased products was relatively uniform regardless of company, brand, or country of origin. In contrast, FBS substitutes obtained from three major livestock species (cattle, pig, and chicken) clearly exhibited differences in composition, a phenomenon that was also observed upon comparing with FBS as well as among different species. Therefore, to replace commercial FBS entirely, the production of a proportionately effective substitute product comprising an equal or similar composition is required, and the results of this study can be a steppingstone to achieving this. In addition, FBS substitutes manufactured using inexpensive slaughter by-products as raw materials are expected to ultimately reduce the unit cost of cultured meat production.

Molecularly imprinted sensor based on poly-o-phenylenediamine-hydroquinone polymer for β-amyloid-42 detection.

A sensitive and selective molecularly imprinted polymer (MIP) sensor was developed for the determination of amyloid-β (1-42) (Aβ42). The glassy carbon electrode (GCE) was successively modified with electrochemical reduction graphene oxide (ERG) and poly(thionine-methylene blue) (PTH-MB). The MIPs were synthesized by electropolymerization with Aβ42 as a template and o-phenylenediamine (o-PD) and hydroquinone (HQ) as functional monomers. Cyclic voltammetry (CV), electrochemical impedance spectroscopy(EIS), chronoamperometry (CC), and differential pulse voltammetry (DPV) were used to study the preparation process of the MIP sensor. The preparation conditions of the sensor were investigated in detail. In optimal experimental conditions, the response current of the sensor was linear in the range of 0.12-10μgmL-1 with a detection limit of 0.018ngmL-1. The MIP-based sensor successfully detected Aβ42 in commercial fetal bovine serum (cFBS) and artificial cerebrospinal fluid (aCSF).

Metagenomic analysis reveals presence of different animal viruses in commercial fetal bovine serum and trypsin.

Animal-derived biological products, such as fetal bovine serum (FBS) and trypsin, are important supplements for scientific, pharmaceutical, and medical use. Although preventive guidelines and tests are implemented to reduce potential viral contamination in these biologicals, they do not target unusual or emerging viruses, leading to safety concerns. Using unbiased metagenomics, we investigated the presence of viruses in recently collected commercial FBS and trypsin samples from different geographic regions. In total, we detected viral sequences belonging to Parvoviridae,Anelloviridae,Flaviviridae, Herpesviridae,Caliciviridae,Nodaviridae,Rhabdoviridae, and Paramyxoviridae, including several viruses related to bovine diseases, viruses of potential human and insect origin, and viruses of unknown origin. Bovine parvovirus 3 and bosavirus were detected with high frequency and abundance in FBS, necessitating more stringent testing for these parvoviruses during production. Both bovine norovirus and bovine viral diarrhea virus 1 displayed relatively high genetic distance to closest hits, indicating the presence of new genotypes in farm animals. While the origin of novel lyssavirus and Nipah virus is unclear, their presence raises the possibility of the introduction of pathogenic animal-derived viruses into biologicals. Our results showed relatively widespread contamination of different viruses in biologicals, underscoring the need for robust safety protocol alternatives, such as metagenomic sequencing, to monitor emerging viruses.

Respiratory signs, fever and lymphopenia in calves inoculated with Brazilian HoBi-like pestiviruses

Hobi-like viruses (HobiPeV) comprise a novel, recently classified species of bovine pestiviruses, originally identified in commercial fetal bovine serum of Brazilian origin and, subsequently, isolated from diseased animals in several countries. Although frequently isolated from clinical cases, most HobiPeV isolates failed to reproduce overt disease in cattle upon experimental inoculation. Herein, we describe the outcome of experimental infection of four to six months-old seronegative calves with two Brazilian HobiPeV isolates. Calves inoculated intranasally with isolate SV478/07 developed viremia between days 2 and 9 post-inoculation (pi) and shed virus in nasal secretions up to day 11pi. These animals presented hyperthermia (day 7 to 10–11 pi) and lymphopenia from days 4 to 8pi. Clinically, all four calves developed varied degrees of apathy, anorexia, mild to moderate respiratory signs (nasal secretion, hyperemia), ocular discharge and pasty diarrhea in the days following virus inoculation. In contrast, calves inoculated with isolate SV757/15 presented only hyperthermia (days 3 to 10–11 pi) and lymphopenia (days 4–8 pi), without other apparent clinical signs. In these animals, viremia was detected up to day 9 pi and virus shedding in nasal secretions lasted up to day 12–14 pi. Both groups seroconverted to the inoculated viruses, developing virus neutralizing (VN) titers from 320 to 5120 at day 28pi. These results extend previous findings that experimental infections of calves with HobiPeV are predominantly mild, yet they also indicate that field isolates may differ in their ability to cause disease in susceptible animals.

Comparison of efficacies of fetal bovine sera from different suppliers in cell culture experiments

Fetal bovine serum (FBS) is widely used as a culture medium supplement. A high degree of variations is observed among FBS samples produced by different suppliers. The purpose of the present study was to compare between the quality and characteristics of FBS produced at Avicenna Research Institute (ARI-FBS) and a commercial FBS. Blood samples were taken from 50 healthy bovine fetuses at 2–9 months of age. Following clotting of the blood, samples were centrifuged. Their sera were collected under sterile conditions. After irradiation, to investigate the performance and quality assurance of the serum samples, some biological properties in cell culture applications, including capacities in supporting single cell growth, proliferation, and cell viability were examined by cell cloning, MTT assay, and flow cytometry analysis. Comparison of ARI-FBS with commercial FBS showed that both of them were expedient and reliable for use in culture studies. However, significant differences in growth rates of different cell lines were observed between them. Results of MTT assay showed that L929 cell line had a significantly better viability in ARI-FBS. Conversely, PC3 and SW480 cell lines were more viable in commercial FBS. In cloning experiments and flow cytometry, both sera showed similar efficiencies. ARI-FBS supported cell growth and maintained cell viabilities in a similar manner to the commercial FBS. ARI-FBS may thus be a proper alternative for commercial FBS as a growth-promoting component for successful culture of cell lines.

Emerging animal viruses: real threats or simple bystanders?

The list of animal viruses has been frequently added of new members raising permanent concerns to virologists and veterinarians. The pathogenic potential and association with disease have been clearly demonstrated for some, but not for all of these emerging viruses. This review describes recent discoveries of animal viruses and their potential relevance for veterinary practice. Dogs were considered refractory to influenza viruses until 2004, when an influenza A virus subtype H3N8 was transmitted from horses and produced severe respiratory disease in racing greyhounds in Florida/USA. The novel virus, named canine influenza virus (CIV), is considered now a separate virus lineage and has spread among urban canine population in the USA. A new pestivirus (Flaviviridae), tentatively called HoBi-like pestivirus, was identified in 2004 in commercial fetal bovine serum from Brazil. Hobi-like viruses are genetically and antigenically related to bovine viral diarrhea virus (BVDV) and induce similar clinical manifestations. These novel viruses seem to be widespread in Brazilian herds and have also been detected in Southeast Asia and Europe. In 2011, a novel mosquito-borne orthobunyavirus, named Schmallenberg virus (SBV), was associated with fever, drop in milk production, abortion and newborn malformation in cattle and sheep in Germany. Subsequently, the virus disseminated over several European countries and currently represents a real treat for animal health. The origin of SBV is still a matter of debate but it may be a reassortant from previous known bunyaviruses Shamonda and Satuperi. Hepatitis E virus (HEV, family Hepeviridae) is a long known agent of human acute hepatitis and in 1997 was first identified in pigs. Current data indicates that swine HEV is spread worldwide, mainly associated with subclinical infection. Two of the four HEV genotypes are zoonotic and may be transmitted between swine and human by contaminated water and undercooked pork meat. The current distribution and impact of HEV infection in swine production are largely unknown. Avian gyrovirus type 2 (AGV2) is a newly described Gyrovirus, family Circoviridae, which was unexpectedly found in sera of poultry suspected to be infected with chicken anemia virus (CAV). AGV2 is closely related to CAV but displays sufficient genomic differences to be classified as a distinct species. AGV2 seems to be distributed in Brazil and also in other countries but its pathogenic role for chickens is still under investigation. Finally, the long time and intensive search for animal relatives of human hepatitis C virus (HCV) has led to the identification of novel hepaciviruses in dogs (canine hepacivirus [CHV]), horses (non-primate hepaciviruses [NPHV] or Theiler's disease associated virus [TDAV]) and rodents. For these, a clear and definitive association with disease is still lacking and only time and investigation will tell whether they are real disease agents or simple spectators.

Complete Genome Sequence of a Bovine Viral Diarrhea Virus 2 from Commercial Fetal Bovine Serum

We isolated a bovine viral diarrhea virus (BVDV) from commercial fetal bovine serum and designated it HLJ-10. The complete genome is 12,284 nucleotides (nt); the open reading frame is 11,694 nt, coding 3,898 amino acids. Phylogenetic analysis indicated that this strain belongs to BVDV group 2.

Methods to select suitable fetal bovine serum for use in quality control assays for the detection of adventitious viruses from biological products

Production of biological products, especially vaccines, usually requires materials derived from animals, and there are always risks that animal pathogens derived from these materials could contaminate the final products. Detection of adventitious agents is performed by quality control tests. In these biological assays, animal derived materials are also used and another problem arises, as fetal bovine serum (FBS) is used as an ingredient in tissue culture media. FBS contaminated with bovine viral diarrhea virus (BVDV) or other bovine pathogens, as well as antibodies against these pathogens may lead to false results in quality control assays. In this study, in order to determine the actual status of commercial FBS, we performed quality tests on various FBS samples. As a result, in 28 of 49 FBS samples (57.1%), pestivirus genes were detected by pan-pestivirus reverse transcription-polymerase chain reaction assay. Furthermore, two samples contained infectious BVDV. Neutralizing antibodies against BVDVs were detected in 48 of 49 samples (97.6%) by the virus neutralization test based on the serum-dilution or virus-dilution methods. Antibodies against other bovine pathogens were detected rarely in these samples. From our results, we recommend methods to select FBS that are focused on detection of BVDV and neutralizing antibodies against BVDV.

Inactivation of Anthracyclines by Serum Heme Proteins

We have previously shown that the anticancer agent doxorubicin undergoes oxidation and inactivation when exposed to myeloperoxidase-containing human leukemia HL-60 cells, or to isolated myeloperoxidase, in the presence of hydrogen peroxide and nitrite. In the current study we report that commercial fetal bovine serum (FBS) alone oxidizes doxorubicin in the presence of hydrogen peroxide and that nitrite accelerates this oxidation. The efficacy of inactivation was dependent on the concentration of serum present; no reaction was observed when hydrogen peroxide or serum was omitted. Peroxidase activity assays, based on oxidation of 3,3',5,5'-tetramethylbenzidine, confirmed the presence of a peroxidase in the sera from several suppliers. The peroxidative activity was contained in the >10000 MW fraction. We also found that hemoglobin, a heme protein likely to be present in commercial FBS, is capable of oxidizing doxorubicin in the presence of hydrogen peroxide and that nitrite further stimulates the reaction. In contrast to intact doxorubicin, the serum + hydrogen peroxide + nitrite treated drug appeared to be nontoxic for PC3 human prostate cancer cells. Together, this study shows that (pseudo)peroxidases present in sera catalyze oxidation of doxorubicin by hydrogen peroxide and that this diminishes the tumoricidal activity of the anthracycline, at least in in vitro settings. Finally, this study also points out that addition of H2O2 to media containing FBS will stimulate peroxidase-type of reactions, which may affect cytotoxic properties of studied compounds.

A infecção pelo vírus da diarréia viral bovina (BVDV) no Brasil: histórico, situação atual e perspectivas

O vírus da Diarréia Viral Bovina (BVDV) possui distribuição mundial e é considerado um dos principais patógenos de bovinos. A infecção e as enfermidades associadas ao BVDV têm sido descritas no Brasil desde os anos 60. Diversos relatos sorológicos, clínico-patológicos e de isolamento do agente demonstram a ampla disseminação da infecção no rebanho bovino brasileiro. Além de sorologia positiva em níveis variáveis em bovinos de corte e leite, anticorpos contra o BVDV têm sido ocasionalmente detectados em suínos, javalis, caprinos, cervos e bubalinos. O BVDV tem sido freqüentemente detectado em fetos abortados, na capa flogística de animais persistentemente infectados (PI) oriundos de rebanhos com problemas reprodutivos, em amostras clínicas e/ou material de necropsia de animais com as mais diversas manifestações clínicas, em sêmen de touros de centrais de inseminação artificial, em fetos saudáveis coletados em matadouros e em soro bovino comercial e/ou cultivos celulares. Aproximadamente 50 isolados do vírus já foram caracterizados genética e/ou antigenicamente, enquanto um número semelhante de amostras aguarda caracterização. A maioria dos isolados caracterizados pertence ao genótipo BVDV-1, biotipo não-citopático (NCP), embora vários isolados de BVDV-2 (e alguns BVDV citopáticos CP) já tenham sido identificados. Os isolados brasileiros apresentam grande variabilidade antigênica, além de diferenças antigênicas marcantes quando comparados a cepas vacinais norte-americanas. Algumas vacinas polivalentes (BHV-1, PI-3, BRSV), contendo o BVDV inativado, têm sido utilizadas no rebanho brasileiro. No entanto, o uso de vacinação ainda é incipiente na maioria das regiões; apenas 2,5 milhões de doses foram comercializadas em 2003. A baixa reatividade sorológica cruzada entre os isolados brasileiros e as cepas vacinais tem estimulado laboratórios nacionais a desenvolver vacinas com isolados autóctones de BVDV-1 e 2. O conhecimento sobre a infecção pelo BVDV no Brasil tem aumentado consideravelmente nos últimos anos, à medida em que cresce o número de laboratórios envolvidos em diagnóstico e pesquisa sobre esse vírus. Diagnóstico sorológico, virológico ou molecular; estudos sobre epidemiologia sorológica e molecular, patogenia e produção de reagentes para diagnósitco têm contribuído para o aumento no conhecimento sobre a infecção pelo BVDV no país.

139 BOVINE AMNIOTIC FLUID FOR THE CULTURE OF TWO-CELL MURINE EMBRYOS

The objective of the experiment was to evaluate bovine amniotic fluid as an alternative medium for culture of two-cell murine embryos. Variability in embryo development rate and high purchase prices of medium supplements such as fetal bovine serum have prompted the search for a serum alternative. Amniotic fluid was collected postmortem from first-trimester bovine fetuses, pooled, heat inactivated at 56°C for 30 min and stored at −20°C until used. Two-cell mouse embryos were collected from (Balb/C × C57BL6) F1 females superovulated with 10 IU PMSG and 10 IU hCG. The experiment was performed to evaluate the effect of commercial fetal bovine serum (F4135, Sigma, South Africa) supplementation to amniotic fluid. Treatments consisted of (1) bovine amniotic fluid, (2) bovine amniotic fluid supplemented with 10% fetal bovine serum, (3) M16 (M7292, Sigma) supplemented with 10% fetal bovine serum, and (4) Medium 199 (M4530, Sigma) supplemented with 10% fetal bovine serum. The latter two media were controls. Twenty-four hours before use the culture media were supplemented with 1% antibiotic antimycotic solution (A5955, Sigma). Media were equilibrated for 24 hours at 37°C and 5% CO2 before use. Embryos were cultured in 50-μL droplets with oil overlay at 37°C in 5% CO2. A minimum of 5 and maximum of 10 embryos per droplet were allowed. Six replications per treatment were done giving a total of 292 embryos in treatment (1), 318 in treatment (2), 304 in treatment (3), and 303 in treatment (4). The embryos were monitored under an inverted microscope (Olympus model IX70) at 24-h intervals for 72 h for blastocyst formation. The differences between embryo growth in the different culture media were assessed by one-way analysis of variance. All culture media supported the development of mouse embryos to the hatched blastocyst stage. A higher (P < 0.05) number of embryos hatched in M16 (64.6%) and Medium 199 (55.0%) supplemented with 10% fetal bovine serum than in frozen bovine amniotic fluid (12.2%) and frozen bovine amniotic fluid supplemented with 10% fetal bovine serum (17.8%). M16 was superior to all other treatments in supporting embryo development up to the morula stage. More than twice the number of embryos (94.9%) reached the morula stage in M16 than in frozen bovine amniotic fluid with (37.4%) or without (29.1%) serum supplementation. Bovine amniotic fluid obtained from postmortem first trimester fetuses supported the development of two-cell mouse embryos; however, embryo development in frozen fetal fluid was lower (P < 0.05) than that obtained in the control media. Fetal bovine serum, when added to amniotic fluid, did not increase the development rate. It seems likely that freezing the amniotic fluid had an adverse effect on in vitro embryo development.

Sensitive method for the determination of bisphenol-A in serum using two systems of high-performance liquid chromatography

The aim of this study was to establish an easy and accurate method for the determination of bisphenol-A (BPA) in the body liquid such as serum and urine. Two high-performance liquid chromatography (HPLC) systems, HPLC with electrochemical detector (ED), and HPLC with mass spectrometry (MS) using electrospray ionization (ESI) interface were used for the assay in the serum samples prepared with solid-phase extraction method. Water or EtOH at a concentration below 50% was suitable for the extraction of BPA from serum. The limit of detection of BPA was 0.2 ng ml −1 for the HPLC-ED method and 0.1 ng ml −1 for HPLC–MS. There was a good correlation between the data obtained by the two HPLC systems. BPA concentrations in healthy human serum were low (0–1.6 ng ml −1). From various commercial fetal bovine serum and sheep plasma, however, significant amounts of BPA were detected. Since no BPA was detected from sheep plasma immediately after collection, the high amounts of BPA were considered to be caused by the handling of blood during the preparation of the products after blood collection. In vitro study showed that the amount of BPA leached from polycarbonate tube into sheep plasma were 40 times larger than those into water and the leached amount of BPA depended on the temperature (37°C>20°C>5°C).

Interaction of nanobacteria with cultured mammalian cells

Nanobacteria were recently isolated from human blood and commercial fetal bovine serum (FBS) and were located in the α-2 subgroup of proteobacteria based upon their 16S rRNA gene sequence. They can be cultured even in the absence of mammalian cells, and have extraordinary properties, like very slow growth rate and an impermeable cell wall, making their detection difficult by standard microbiological techniques. Since they are present in FBS, and thus in cell cultures, it is essential to clarify their effects on cultured mammalian cells. In this study, we show that four out of six nanobacterial isolates from different sera exerted a cytotoxic effect on 3T6 fibroblasts verified by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] viability assay, lactate dehygrogenase (LDH) release and by direct microscopy. The cytotoxic effect of nanobacteria was attenuated after they had been subcultured several times. The cytotoxic effect was similar with all tested murine and human fibroblastoid cell lines. Differential interference contrast and electron microscopy, and FITC staining with specific monoclonal antibodies indicated selective, possibly receptor-mediated adherence, followed by internalization and cytotoxicity in the 3T6 fibroblasts used as a model in these interaction studies. Thus, nanobacteria have a special way of invading mammalian cells: they trigger cells that are not normally phagocytic to engulf them. These organisms seem to be an important cause for cell vacuolization, poor thriving and unexpected cell lysis, problems frequently encountered in mammalian cell culture.

Supplementation of HAM's F10 culture medium with three different sera in the culturing of baboon oocytes

1. 1. Ten female baboons ( Papio ursinus) were stimulated for a total of 20 cycles with 3 ovulation induction agents. 2. 2. Oocytes obtained were randomly allocated to Ham's F10 culture medium supplemented with human fetal cord serum, primate serum or commercial fetal bovine serum respectively. 3. 3. Fertilization occured (38.1–45.5%) in all 3 supplements, but cleavage and embryo development was more successful in culture medium supplemented with fetal bovine serum. 4. 4. Eight embryos were cultured and 6 (75%) of these were cultured in fetal bovine serum supplemented medium.

Role of serum in survival of Treponema pallidum in tissue culture.

During attempts to cultivate Treponema pallidum, it was determined that length of time for survival of virulent treponemes was highly dependent on the quality of the fetal bovine serum (FBS) used as a protein supplement in the culture medium. Eighteen lots of commercial FBS were tested for their ability to maintain survival of T. pallidum in cultures of cottontail rabbit epithelial (SflEp) cells. All were capable of supporting growth of these cells. However, in tests on five of the lots, attachment of treponemes to the SflEp cells was either extremely poor or the 50% survival time (ST50) was less than 5 days. With two of these lots, no treponemes survived for 5 days. By contrast, in tests with 11 of the FBS lots, the ST50 of the treponemes was 12 days or greater; however, there was a great variation in the number of treponemes that attached. Selection of lots of FBS for ultimate experimental use was based on their influence both to extend length of time for survival of treponemes and to increase the number of treponemes that attached to the SflEp cells during that period.

Acholeplasma morum, a New Non-Sterol-Requiring Species

Two acholeplasmas recovered from contaminated cell cultures or commercial fetal bovine serum were found to be similar in biochemical and serological properties. The organisms, although not requiring serum or cholesterol for growth, did require a supplement of 0.5% albumin and 10 μg of palmitic acid per ml to a serum-free medium for sustained growth in the absence of sterols. The growth of both organisms was inhibited in medium containing 20% horse serum, although smaller amounts of horse serum or 20% fetal bovine serum did not exert inhibitory effects. The two strains possessed biochemical properties typical of other non-sterol-requiring members of the Mycoplasmatales, and they were serologically distinct from seven established Acholeplasma species. On the basis of these findings and other morphological, biological, and serological properties of the organisms, it is proposed that these organisms represent a new species, Acholeplasma morum. Strain 72–043 (=ATCC 33211) is the type strain.

Detection of antibody to bovine syncytial virus and respiratory syncytial virus in bovine fetal serum.

Batches of commercial fetal bovine serum, described by the suppliers as antibody-free, all contained antibody to bovine syncytial virus (BSV) when tested by indirect immunofluorescence. Antibody to bovine respiratory syncytial virus (RSV) was not detected in these sera. Twenty-four percent of individual fetal bovine sera contained antibody to BSV, and 14% contained antibody to RSV when tested by indirect immunofluorescence. BSV antibody titers in fetal sera from dams with high BSV antibody levels were variable but always higher than RSV antibody titers. Radial immunodiffusion studies with BSV-positive sera revealed the presence of immunoglobulin M (IgM), IgG, and IgA, but the quantity of these immunoglobulins was not directly related to the BSV antibody titers. The evidence suggests that the antibody present in fetal sera arose as the result of infection rather than from maternal transfer across the placenta.