Antiophidian sera sterility control: topics in perspective / Controle de esterilidade de soros antiofídicos: temas em perspectiva

The aim of this work is to review the most important topics about the antiophidic sera sterility, including obtaining methods, sterilization procedures and clean room control using Vital Brazil Institute (VBI) as an example. Bibliographical research was performed through Medline, Lilacs, PubMed, ISI and the Fundação Oswaldo Cruz - RJ and VBI Libraries, from 1960 to 2009. The antiophidic sera for human use are immunobiologic products produced in Brazil by three national laboratories, including VBI. Due to the parenteral use, these products should be sterile and pyrogen-free, which demands the microbiological control during the whole fabrication process. The sterility and pyrogen tests are important steps to ensure the quality and safety of these immunobiological products. Thus, these tests are target for continue evaluation and improvement. The most interfering aspects in the consistency and analytical patterns include the proper method selection, sampling, culture conditions and validation criteria. As the national and international legal requirements are cautious with the assays validation and approval of sterile parenteral products; the intrinsic limitations for established assays still require more investigation aiming the continue improvement of the microorganism and contaminants detection methods and optimization of the analysis extent.

O objetivo deste trabalho é revisar os tópicos mais relevantes para o controle da esterilidade de soros antiofídicos, abordando-se métodos de obtenção, procedimentos de esterilização e o controle de áreas limpas utilizando como exemplo os procedimentos adotados pelo Instituto Vital Brazil (IVB). Um levantamento bibliográfico foi realizado no Medline, ISI, Biblioteca da Fundação Oswaldo Cruz-RJ e IVB, no período de 1960 a 2009. Os soros antiofídicos para uso humano são produtos imunobiológicos fabricados no Brasil por três laboratórios nacionais, dentre eles o IVB. Por serem de administração parenteral, devem ser obrigatoriamente estéreis e apirogênicos, exigindo controle microbiológico durante todo o processo de fabricação. O teste de esterilidade e apirogenia são importantes instrumentos para garantir a qualidade e segurança microbiológica desses produtos, sendo alvo de avaliações constantes para seus aprimoramentos. Os aspectos que mais interferem na sua consistência e valor analítico incluem a escolha adequada do método, amostragem, condições de cultivo e critérios de validação. À medida que os requisitos legais nacionais e internacionais se mostram rigorosos na validação de ensaios e aprovação de produtos estéreis parenterais, limitações intrínsecas ao ensaio padronizado requisitam mais investigações, objetivando o aprimoramento contínuo nos métodos de detecção de microorganisms, contaminantes e otimização do tempo total de análise.

Imunologia reprodutiva na prática clínica: uma visão crítica / Reproductive immunology in clinical practice: a critical view

Atualmente, grande importância tem sido dada aos aspectos imunológicos da reprodução humana. Embora a maior parte dos mecanismos imunológicos envolvidos na gestação não seja inteiramente compreendida, muitos casais inférteis têm sido submetidos à investigação imunológica e diferentes tipos dos assim chamados tratamentos imunológicos têm sido amplamente prescritos. Os exames que devem ser incluídos nesta investigação, bem como a eficácia da maioria dos tratamentos ainda não estão bem estabelecidos. O objetivo deste artigo é discutir o que se sabe sobre o real papel do sistema imune em casos de infertilidade e apresentar uma visão crítica da imunologia reprodutiva na prática clínica.

Anti-zona pellucida antibodies in infertile patients in relation to multiple puncture of ovaries and unexplained infertility

Auto antibodies to zona-pellucida [AZA] seem to be important autoantibodies implicated in reproduction, with substantial role in both endocrine and reproductive functions of the human ovary. There are some debates on the relation of AZA with infertility, repeated In Vitro Fertilization [IVF] attempts, and outcome of it. In this study, we assessed the presence of AZA in the follicular fluids [FFs] of women who underwent intra cytoplasmic sperm injection [ICSI], in relation to etiology of infertility and multiple puncture of ovaries. In this prospective study, follicular fluids were evaluated from 96 infertile women, [19-40 years old, 31.5 +/- 5.1], who were c and idates for ICSI based on the etiology of infertility. From these 80 women had explained infertility whereas 16 had unexplained infertility. All FFs were evaluated for presence of AZA by ELISA test. Twenty patients [20.8%] were positive for AZA in follicular fluid. In patients with unexplained infertility, AZA antibody in follicular fluid, was significantly higher than the group with proven etiology of infertility [p=0.001]. In addition, 20.4% of patients who had been punctured previously showed AZA in their FFs which is statistically similar to the patients who were punctured for the first time. The high incidence of AZA in infertile women, especially women with unexplained infertility has to be considered. Relation of the presence AZA and repeated puncture of ovaries is still debatable. Determinations of AZA are highly recommended in evaluation of infertile couples especially in patient with unexplained infertility

[Designing an ELISA method using sperm antigens for detection of antisperm antibody in comparison with SpermMar test

The role of antisperm antibodies with a prevalence of 6-26% is well known in immunological infertility. Thus, there is clinical importance to determine ASA levels in both male and female. Nowadays, one of the most important discussed controversies in the field of immunological infertility is establishing an standard method to determine ASA. It seems that ELISA method will be more sensitive, specific and more diagnostic in determination of ASA if sperm surface antigens could be used as coated antigens, with least contamination to sperm intracellular antigens and nonspermic antigens. So, the aim of this study is designing an ELISA method by using the best method of sperm antigens extraction with at least contamination. In this study we designed an ELISA method with three different extraction methods of sperm antigens including sonication method, using SDS detergent, and application of LIS detergent, then we compared ELISA method based on the three extraction methods as well as two similar commercial ELISA kit [IBL Co, and Bioserv Co] with SpermMar test. Comparing designed method with commercial kit indicated that among 28 sera which had 16 positive sera and 12 negative sera by SpermMar, 14 sera were true positive by LIS method and only 2 cases were false negative without any false positive results, whereas there were 5 true positive results and 11 cases false negative by the sonication method. The SDS method also had 13 true positive results with 3 false negative and 4 false positive results. In addition, two commercial kit had in turn 7 and 4 cases true positive and both of them had 1 case false positive and in turn 9 and 12 cases with false negative result. ELISA method designed by LIS detergent has adequate sensitivity [87.5%] with higher specificity [100%] and efficacy [92.8%] than other extraction methods. There is a significant correlation between this designed method and SpermMar test [r=0.572]. The results of this study indicated that ELISA method by LIS antigens has at least contamination with nonspermic antigens and it is better than other extraction methods and commercial ELISA kits for detection of antisperm antibody

Immunohistochemical study of HLA-DR antigen in endometrial tissue of patients with endometriosis / 华中科技大学学报（医学）（英德文版）

In order to evaluate the expression of HLA-DR antigen in glandular cells in eutopic and ectopic endometrium in patients with endometriosis, 19 infertile patients with endometriosis were analyzed immunohistochemically by labelled streptavidin biotin (LSAB) method. Nineteen infertile patients without endometriosis were studied as controls. The results showed that the expression of HLA-DR antigen in the glandular cells in both eutopic and ectopic endometrium was increased significantly as compared with that in the controls (P < 0.01). It is likely that aberrant expression of HLA-DR antigen in endometriotic tissue is involved in abnormal immunogenesis of endometriosis.

Anticuerpos antiespermatozoides en parejas con infertilidad de causa no explicada / Antisperm antibodies in couples with infertility of unknown origin

Se estudió una muestra de 75 parejas infértiles en relación con la presencia de anticuerpos antiespermatozoides en plasma seminal, moco cervical o adheridos a los espermatozoides, mediante la prueba de inmunobeads y se determinó la capacidad inhibidora del plasma seminal sobre la proliferación linfocitaria, para evaluar la capacidad inmunomoduladora del semen como participante en el mecanismo patogénico primario de aparición de los anticuerpos antiespermatozoides. Se organizaron las parejas en 2 grupos, según la presencia o no de anticuerpos antiespermatozoides y se compararon en cuanto a la frecuencia de embarazo y la capacidad inmunosupresora del plasma seminal. Además se comparó la incidencia de embarazo en grupos de parejas con capacidad inmunosupresora y sin ella. Se observó que la presencia de anticuerpos antiespermatozoides fue menos frecuente que la reportada por otros autores y no estuvo relacionada con la capacidad inmunosupresora del plasma seminal, tampoco influyeron sobre la frecuencia de embarazo la presencia de anticuerpos antiespermatozoides y la capacidad inmunosupresora del plasma seminal(AU)

Immunological assessment of infertility by estimation of antisperm antibodies in infertile couples.

160 clinical samples were collected from 40 infertile couples with unexplained infertility. The samples collected included serum and seminal plasma of the male partners and serum and cervical mucus samples of the female partners. 25 fertile healthy couples were investigated as controls. All the samples collected were then tested for class-specific antisperm antibodies by an Enzyme linked immunosorbent assay (ELISA). Antisperm antibodies were detected in 30% of the infertile couples which included 25% female and 10% male partners. Amongst the cases positive for antisperm antibodies, antibodies were detected most frequently in female sera 58.4% followed by male sera 33% and 25% in cervical mucus. The isotyping of antisperm antibodies in various samples showed IgG to be the most frequent type specific antibody followed by IgM & IgA types of antibodies. ELISA has provided a relatively simple, reliable and highly reproducible method of detection of antisperm antibodies. Thus application of antisperm antibody testing especially in cervical mucus should become an integral part of the investigation of immunologic infertility.

Aspectos imunológicos da infertilidade conjugal / Immunological aspects of human infertility

Na infertilidade conjugal, apesar do avanço tecnológico, nao houve grande melhora nas taxas de gestaçao à termo. Uma lacuna existe no conhecimento sobre os mecanismos inerentes ao processo reprodutivo. Assim, atençao especial tem sido dada ao papel do sistema imunológico na reproduçao, uma vez que o feto deverá suscitar uma resposta imune no organismo materno. Os fatores que seriam responsáveis pela manutençao da gestaçao continuam sendo investigados. Apesar da fisiologia deste processo nao ser bem conhecida, alguns métodos diagnósticos têm sido empregados com o intuito de melhorar o prognóstico destes casais. Estes métodos trouxeram algumas contribuiçoes, mas há de se dizer que o processo reprodutivo é um sistema biológico complexo, que envolve o equilíbrio de vários sistemas orgânicos. Esta revisao procura mostrar aspectos atuais sobre este tema.

Factores embriotóxicos tempranos como posible mecanismo de infertilidad en pacientes con aborto habitual y esterilidad sin causa aparente / Embryologic toxic factors as infertility cause in patients habitual abortions with sterility without apparent cause

El objetivo del presente estudio es identificar la presencia de factores embriotóxicos tempranos como causa de infertilidad. Se estudia 15 sueros de pacientes divididos en dos grupos: de abortos habituales y con esterilidad sin causa aparente. Se realizó cultivo de embriones preimplantacionales de ratón que permite identificar en el suero de mujeres agentes embriotóxicos y mutagénicos incompatibles con el desarrollo embrionario.

Infertilidad inmunológica / Immunological infertility

Hasta el 30 por ciento de las infertilidades inexplicadas, tienen origen inmunológico. La infertilidad inmunológica agrupa una serie de trastornos en los que solamente ahora, se puede contar con la tecnología adecuada para diagnosticarla. Esta revisión comprende la fisiopatología de la formación de los anticuerpos anti-esperma, tanto en el hombre como en la mujer, y las técnicas para hacer un mejor enfoque diagnóstico de esta compleja enfermedad, sobre todo en nuestro medio, haciendo énfasis en la prueba de penetración del moco cervical in vitro. También se revisan los tratamientos que en la actualidad ofrece la medicina...

Circulating iso and auto antibodies to human spermatozoa in infertility.

Blood samples from 50 couples of unexplained primary infertility and 25 healthy fertile couples were tested for antispermatozoa antibodies (ASA) by gelatin agglutination test and sperm immobilising complement dependent cytotoxicity test. The incidence of sperm agglutinating and sperm immobilising antibodies was found to be 14 per cent and 10 per cent respectively in infertile couples. Agglutinating antibodies were detected in 18 per cent males and 16 per cent of females while cytotoxic antibodies were found in 14 per cent males and 10 per cent of females. Only 4 per cent of the normal fertile couples taken as a control group showed positive ASA, hence the difference in incidence of positive ASA between the infertile and fertile couples was found to be statistically significant. Also, the agglutination test was observed to be more sensitive than the cytotoxicity test.

Anticuerpos ABH en moco cervical / Antybodies ABH in cervical mocus

Se estudian anticuerpos anti-A y anti-B en moco cervical pre y postratamiento con solución de 2-mercaptoetanol, utilizado para romper puentes disulfuro intracatenario de los anticuerpos IgM

Immunological aspects of infertility in cows and buffaloes in Egypt

The present investigation revealed that the immunological reaction between the cervical mucus of repeat breeder cow and washedl spermatozoa gave a higher titre. On the other hand the reaction of the cervical mucus of normal cow and buffalo-cow with the seminal plasma gave a low agglutinating titre. The influence of the cervical mucus of repeat breeder cow- and buffalo-cow on the homologus bull semen gave a higher reaction than that with the heterologous bull semen. The results were attributed to the presence of agglutinating and immobilizing antibodies against the semen in the cervical mucus of the repeat breeder cows and buffalo-cows

Anticuerpos antiespermatozoide en la infertilidad / Anti-spermatozoa antibodies in infertility

Utilizando la tecnica de macro-aglutinacion en gelatina propuesta por Kibrick, se estudio la presencia de anticuerpos antiespermatozoide en parejas infertiles. De 52 parejas, 33 consultaron por infertilidad primaria (63.6%) y 19 por secundaria (36.5%). El 46.1% de todas estas mujeres presentaron anticuerpos circulantes a titulos de 1:16 a 1:128, contra solo 3.8% en el grupo control. La frecuencia de anticuerpos circulantes por infertilidad primaria que en aquellas que lo hicieron por infertilidad secundaria (36.8). Solo 12 hombres dentro del grupo estudiado, presentaron anticuerpos a titulos menores o iguales a 1:16; no hubo en ellos correlacion ninguna con la calidad de los espermatozoides. No se encontro ninguna relacion entre la patologia gineco-obstetrica y la presencia de los anticuerpos circulantes