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A periodontite é caracterizada pela inflamação nos tecidos periodontais e consequente perda das estruturas de suporte do dente: osso alveolar, cemento e ligamento periodontal, podendo levar à perda de dentes. A homeostase do tecido ósseo é gerida fundamentalmente pela regulação da via de sinalização composta pelo Receptor Ativador do fator Nuclear B (RANK), seu ativador RANK Ligante (RANKL), e o falso ligante de RANKL Osteoprotegerina (OPG). O processo inflamatório destrutivo e com gatilho incerto da doença periodontal aumenta a expressão de RANKL, levando a desregulação no turnover ósseo, desbalanceado para reabsorção maior que a formação do osso alveolar. Visando reduzir danos ou promover resolução da inflamação, terapias de modulação da resposta imune do hospedeiro surgem como alternativa para manuseio da periodontite para além da abordagem sobre microrganismos uma vez que indivíduos respondem de maneiras diferetenes ao tratamento e possuem diferentes padrões de perda óssea. Anticorpos anti-RANKL demonstraram serem capazes de diminuir a reabsorção óssea em processos inflamatórios e já foram apontados com potencial terapêutico sobre a patologias que levam a reabsorção óssea, bem como a periodontite. Com o propósito de verificar o potencial de modulação da perda óssea alveolar por anticorpo anti-RANKL sobre a doença periodontal induzida em animais, este estudo foi conduzido seguindo as normas PRISMA e da Cochrane Lybrary. Estudos pré-clínicos em modelo animal de periodontite experimental foram considerados elegíveis. A busca eletrônica incluiu as bases de dados MEDLINE, EMBASE e LILACS e Web of Science para artigos publicados até agosto de 2022. O Risco de Viés foi analisado através da ferramenta Systematic Review Center for Laboratory Animal Experimentations. Foram incluídos 5 de 326 estudos encontrados nas bases de dados. Anticorpos anti-RANKL de diferentes origens foram ministrados em diferentes modelo de periodontite experimental induzida. Os estudos selecionados apresentaram importantes características a serem observadas na condução de estudos em animais. Essa revisão sistemática concluiu que a modulação da interação entre RANK e RANKL mediada por anticorpo anti-RANKL apresenta um potencial terapêutico adjunto ao manuseio da doença periodontal desde que considerado o momento de uso dessa abordagem. Além disso, mais estudos de farmacodinâmica e farmacocinética são necessários para aplicações sobre a periodontite.

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Inflammatory myopathies belong to a heterogeneous group of autoimmune diseases, which are mainly characterized by muscle weakness, elevated muscle enzymes, myopathic changes on electromyography, and muscle biopsy abnormalities. They may also have extraocular manifestations as well as involvement of the lungs, skin, and joints. This descriptive study analyzes cases of idiopathic inflammatory myopathies (IIM) diagnosed and treated at the Regional Hospital of Talca between 2016 and 2021 where data on clinical presentation and serological profile. Nineteen cases were identified, mainly dermatomyositis, with a higher proportion of women affected. Most patients had elevated antinuclear antibodies and the most frequently found antibodies were RO52 and JO1. These results provide valuable information on IIM in the Maule region and may contribute to improving the diagnosis and management of these diseases.

Las miopatías inflamatorias pertenecen a un grupo heterogéneo de enfermedades autoinmunes, que se caracterizan principalmente por debilidad muscular, elevación de enzimas musculares, cambios miopáticos en la electromiografía y anomalías en la biopsia muscular. Pueden además tener manifestaciones extraoculares como también afectación de los pulmones, piel, y articulaciones. Este estudio descriptivo analiza casos de Miopatías inflamatorias idiopáticas (MII) diagnosticados y tratados en el Hospital Regional de Talca entre 2016 y 2021 donde se recopilaron datos sobre presentación clínica y perfil serológico. Se identificaron 19 casos, principalmente dermatomiositis, observándose una mayor proporción de mujeres afectadas. La mayoría de los pacientes presentó anticuerpos antinucleares elevados y los anticuerpos más frecuentemente encontrados fueron el RO52 y JO1. Estos resultados proporcionan información valiosa sobre las MII en la región del Maule y pueden contribuir a mejorar el diagnóstico y manejo de estas enfermedades.

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A aloimunização é caracterizada pela produção de anticorpos na vigência de ex- posição a antígenos não próprios, estando mais relacionada ao sistema ABO/Rh. Contudo, infrequentemente, a doença pode surgir por meio da exposição por antí- genos de outros sistemas sanguíneos, como o sistema MNSs. O caso deste estudo relata o acompanhamento do pré-natal de uma paciente com tipagem sanguínea O+ e Coombs indireto reagente com variação de titulação ao longo das consultas, sendo evidenciada a presença de anti-M IgM por pesquisa de anticorpos antieri- trocitários irregulares, entretanto sem repercussões fetais. Com isso, no decorrer das análises bibliográficas, notou-se que o Coombs indireto (antiglobulina hu- mana anti-IgG) é capaz de reagir com as imunoglobulinas A e M, sendo, portanto, teoricamente capaz de ser positivado pelo anti-M IgM. Com isso, e devido ao fato de o fator IgM não atravessar a barreira transplacentária, não houve complicações fetais no presente caso clínico.

Alloimmunization is characterized by the production of antibodies in the presence of exposure to non-self antigens, most commonly associated with the ABO/Rh system. However, infrequently, the condition can arise from exposure to antigens from other blood systems, such as the MNSs system. This study reports the prenatal monitoring of a patient with blood type O+ and a reactive indirect Coombs test with titration variations during consultations. The presence of anti-M IgM was evidenced through irregular anti-erythrocyte antibody screening; however, no fetal repercussions were observed. Throughout the bibliographic analyses, it was observed that the indirect Coombs test (human anti-IgG globulin) is capable of reacting with immunoglobulins A and M. Therefore, it is theoretically possible to be positive due to anti-M IgM. Con- sequently, as the IgM factor does not cross the transplacental barrier, there were no fetal complications in the present clinical case.

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A resposta imunológica pelo SARS-CoV-2 após protocolos vacinais e infecção natural é pouco compreendida. Comparando indivíduos vacinados com esquema heterólogo que receberam um reforço vacinal (imunidade vacinal) com aqueles que apresentaram episódio leve de COVID-19 (imunidade híbrida) no mesmo período, verificamos níveis semelhantes de anticorpos contra SARS-CoV-2. Em culturas de células mononucleares, o estímulo com o antígeno viral induziu produção de citocinas pró-inflamatórias nos dois grupos, entretanto, os níveis de IL-17 foram menores em indivíduos com imunidade vacinal. Nossos resultados sugerem que o reforço vacinal teve efeitos semelhantes à infecção natural pelo SARS-CoV-2 na resposta imunológica de indivíduos previamente vacinados. (AU)

The immune response generated by SARS-CoV-2 vaccination protocols and natural infection remains incompletely understood. We compared individuals who received a heterologous vaccination scheme with a booster shot (vaccine immunity) to those who experienced a mild COVID-19 episode (hybrid immunity) during the same timeframe. Our findings revealed similar levels of SARS-CoV-2 antibodies in both groups. Stimulation by viral antigen in mononuclear cell cultures induced pro-inflammatory cytokines in both groups, while individuals with vaccine immunity exhibited lower IL-17. These results suggest that a vaccine booster can induce an immune response in previously vaccinated individuals comparable to that elicited by natural SARS-CoV-2 infection. (AU)

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OBJECTIVE@#To explore effect of nerve growth factor (NGF) antibody on knee osteoarthritis (KOA) pain model was evaluated by in vitro model.@*METHODS@#Thirty male SPF rats aged 28-week-old were divided into blank group (10 rats with anesthesia only). The other 20 rats were with monoiodoacetate (MIA) on the right knee joint to establish pain model of OA, and were randomly divided into control group (injected intraperitoneal injection of normal saline) and treatment group (injected anti-NGF) intraperitoneal after successful modeling, and 10 rats in each group. All rats were received retrograde injection of fluorogold (FG) into the right knee joint. Gait was assessed using catwalk gait analysis system before treatment, 1 and 2 weeks after treatment. Three weeks after treatment, right dorsal root ganglia (DRG) were excised on L4-L6 level, immunostained for calcitonin gene-related peptide (CGRP), and the number of DRGS was counted.@*RESULTS@#In terms of gait analysis using cat track system, duty cycle, swing speed and print area ratio in control and treatment group were significantly reduced compared with blank group (P<0.05). Compared with control group, duty cycle and swing speed of treatment group were significantly improved (P<0.05), and there was no significant difference in print area ratio between treatment group and blank group (P>0.05). The number of FG-labeled DRG neurons in control group was significantly higher than that in treatment group and blank group (P<0.05). The expression of CGRP in control group was up-regulated, and differences were statistically significant compared with treatment group (P<0.05).@*CONCLUSION@#Intraperitoneal injection of anti-NGF antibody inhibited gait injury and upregulation of CGRP in DRG neurons. The results suggest that anti-nerve growth factor therapy may be of value in treating knee pain. NGF may be an important target for the treatment of knee OA pain.

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Plant bioreactor is a new production platform for expression of recombinant protein, which is one of the cores of molecular farming. In this study, the anti DYKDDDDK (FLAG) antibody was recombinantly expressed in tobacco (Nicotiana benthamiana) and purified. FLAG antibody with high affinity was obtained after immunizing mice for several times and its sequence was determined. Based on this, virus vectors expressing heavy chain (HC) and light chain (LC) inoculated into Nicotiana benthamiana leaves by using Agrobacterium-mediated delivery. Accumulation of the HC and LC was analyzed by SDS/PAGE followed by Western blotting probed with specific antibodies from 2 to 9 days postinfiltration (dpi). Accumulation of the FLAG antibody displayed at 3 dpi, and reached a maximum at 5 dpi. It was estimated that 66 mg of antibody per kilogram of fresh leaves could be obtained. After separation and purification, the antibody was concentrated to 1 mg/mL. The 1:10 000 diluted antibody can probe with 1 ng/mL FLAG fused antigen well, indicating the high affinity of the FLAG antibody produced in plants. In conclusion, the plant bioreactor is able to produce high affinity FLAG antibodies, with the characteristics of simplicity, low cost and highly added value, which contains enormous potential for the rapid and abundant biosynthesis of antibodies.

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Introducción: en la esclerosis sistémica (ES) la positividad de los anticuerpos es útil para el diagnóstico y la clasificación siendo generalmente autoexcluyentes. Objetivos: estimar la prevalencia de anticuerpos anti-RNA polimerasa III (anti-RNAP III) positivos en pacientes con anti-topoisomerasa I (ATA) y anticentrómero (ACA) negativos, y describir las características clínicas. Materiales y métodos: estudio bidireccional, descriptivo, unicéntrico. Se incluyeron pacientes mayores de 18 años que cumplían criterios clasificatorios del American College of Rheumatology/European League Against Rheumatism (ACR/EULAR) 2013, con ATA y ACA negativos. Se determinó anti-RNAP III por ELISA, QUANTA lite RNA POL III Inova Diagnostics®. Se compararon los pacientes anti-RNAP III positivos versus anti-RNAP III negativos. Resultados: se incluyeron 31 pacientes. El 48,4% (15/31) anti-RNAP III positivo y el 51,6% (16/31) negativo. La prevalencia de anti RNAP-III positivo fue de 5,06%. Se observó mayor proporción de úlceras digitales (40% versus 25%), amputaciones (13,3% versus 6,3%), artritis (33% versus 18,8%), frote tendinoso (20% versus 7,1%) y ectasia vascular antral gástrica (EVAG, 16,7% versus 0%) en el grupo anti-RNAP III positivo. Conclusiones: debido a su posible rol como marcador pronóstico, resulta fundamental conocer las características de nuestra población y, en especial, de nuestro hospital(AU)

Introduction: in systemic sclerosis (SS), autoantibody positivity is useful for diagnosis and classification, being mutually self-exclusive. Objectives: estimate the prevalence of positive anti-RNA polymerase III (anti-RNAP III) antibodies in patients with negative anti-topoisomerase I (ATA) and anti-centromere (ACA) and describe the clinical characteristics. Materials and methods: bidirectional, descriptive, single-center study. We included patients over 18 years of age who met the American College of Rheumatology and European League Against Rheumatism (ACR/EULAR) 2013 classification criteria with ATA and ACA negative. anti-RNAP III was determined by ELISA, QUANTA lite RNA POL III Inova Diagnostics®. Anti-RNAP III positive patients were compared with anti-RNAP III negative patients. Results: 31 patients were included. 48.4% (15/31) anti-RNAP III positive and 51.6% (16/31) negative. The prevalence of positive anti RNAP-III was 5.06%. A higher proportion of digital ulcers (40% vs 25%), amputations (13.3% vs 6.3%), arthritis (33% vs 18.8%), tendon rub (20% vs 7.1%) and gastric antral vascular ectasia (GAVE) were observed (16.7% vs 0%) in the anti-RNAP III positive group. Conclusions: due to its possible role as a prognostic marker, it seems essential to know the characteristics of our population and especially of our hospital(AU)

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O conhecimento sobre a durabilidade da resposta imune mediada pela infecção natural ou pela vacina contra a COVID-19 é fundamental para a proteção contra novas infecções por SARS-CoV-2. Anticorpos neutralizantes são indicadores importantes de imunidade, mas altos títulos têm sido associados a casos graves. A exposição prolongada aos antígenos promove a hipermutação somática, essencial para a maturação da afinidade dos anticorpos; no entanto, a infecção por SARS-CoV-2 geralmente gera anticorpos de afinidade baixa a intermediária, e ainda há debate sobre o número de doses necessárias na vacinação para alcançar maturação completa e imunidade protetora. Para avaliar o perfil da resposta humoral contra o SARS-CoV-2, realizou-se um acompanhamento longitudinal de profissionais da saúde, analisando a produção, prevalência e maturação da afinidade da imunoglobulina G (IgG) anti o domínio de ligação do vírus (RBD). Amostras de soro de indivíduos com histórico de COVID-19 (CoV) e sem infecção documentada (nCoV), coletadas antes e após a vacinação (duas doses de CoronaVac [CN] ou ChAdOx/AstraZeneca [AZ], seguidas de uma dose de reforço com BNT162b2 [PF]), foram avaliadas por ensaios Imunoenzimáticos (ELISA). Os resultados foram expressos como Índice ELISA (IE), Índice de Avidez (IA) e títulos de anticorpos neutralizantes (NAbs). Dos 297 voluntários imunizados, a maioria (90%) recebeu a CN. Após a 2ª dose (35 dias), 88% apresentaram anticorpos IgG anti-RBD...(AU)

Understanding the durability of the immune response mediated by natural infection or vaccination against COVID-19 is fundamental for protection against new SARS-CoV-2 infections. Neutralizing antibodies are important indicators of immunity, but high titers have been associated with severe cases. Prolonged exposure to antigens promotes somatic hypermutation, essential for antibody affinity maturation; however, SARS-CoV-2 infection generally generates antibodies with low to intermediate affinity, and there is still debate about the number of doses required in vaccination to achieve complete maturation and protective immunity. To assess the profile of the humoral response against SARS-CoV-2, a longitudinal follow-up of healthcare professionals was conducted, analyzing the production, prevalence, and affinity maturation of immunoglobulin G (IgG) against the virus receptor binding domain (RBD). Serum samples from individuals with a history of COVID-19 (CoV) and without documented infection (nCoV), collected before and after vaccination (two doses of CoronaVac [CN] or ChAdOx/AstraZeneca [AZ], followed by a booster dose with BNT162b2 [PF]), were evaluated by...(AU)

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Background and Aims: Some studies have reported the coexistence of inflammatory bowel disease (IBD) and celiac disease (CD). However, the prevalence of anti-tissue transglutaminase antibodies (IgA and IgG) and their screening value in patients with IBD is not yet clear. This study aimed to assess the prevalence of IgA anti-tTG and its potential correlation with disease status in patients with IBD. Materials and Methods: This cross-sectional study was conducted on 110 patients with confirmed IBD diagnosis at Ghaem Hospital, Mashhad, Iran. For each patient, all demographic and clinical data including age, extra intestinal manifestations, underlying diseases, types of diseases, and surgical history were collected. IgA anti-tissue transglutaminase titers were assessed by enzyme-linked immunosorbent assay. Results: None of the patients with IBD were positive for IgA anti-tTG antibodies, with a mean titer of 3.31 ± 1.3 AU/mL. Also, the mean titers were not associated with age, gender and various disease clinical features including the disease history, underlying disease, diagnosis type, extraintestinal manifestations, and surgery history. Conclusion: No significant prevalence pattern of IgA anti-tTG antibody was observed in patients with IBD. Accordingly, serological screening for CeD is not recommended in IBD patients, unless in a relevant clinical CeD suspicion. (AU)

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INTRODUCCIÓN. La Encefalitis Autoinmune es una afección inflamatoria severa del sistema nervioso central mediada por anticuerpos. Su diagnóstico en pediatría es complejo, por lo que ahondar en su cuadro clínico, métodos diagnósticos y tratamiento es relevante. OBJETIVO. Establecer el cuadro clínico, el diagnóstico y el tratamiento de la Encefalitis Autoinmune en pacientes menores de 18 años. MATERIALES Y MÉTODOS. Revisión sistemática. A través de las bases PubMed, Google Scholar, The Lancet se realizó la búsqueda de artículos publicados en idioma inglés y español (2016-2021), con las siguientes palabras clave: encefalitis autoinmune AND pediatría OR niños AND diagnóstico AND anticuerpos OR tratamiento OR inmunoterapia. La calidad de los estudios se evaluó con la escala NIH Quality Assessment Tools. La heterogeneidad de los datos únicamente permitió el calculó de las frecuencias de sintomatología, y se hizo una descripción narrativa de los hallazgos. RESULTADOS. De un total de 100 artículos, 15 fueron seleccionados. La encefalitis por anticuerpos contra el receptor de N-metil-D-aspartato es la frecuente. Las convulsiones (52.6%), los trastornos del movimiento (45%) y los cambios de la personalidad o conducta (44.4%) forman la sintomatología de presentación clínica de esta afección más frecuente en los niños. El estudio de líquido cefalorraquídeo presentó pleocitosis linfocitaria y aumento de proteínas. El tratamiento con inmunoterapia empírica luego de descartar causas infecciosas o metabólicas es seguro, en comparación a ningún tratamiento. CONCLUSIÓN. La encefalitis autoinmune contra receptor de N-metil-D-aspartato es más frecuente en niños, tiene diversidad de presentación clínica, y mejora con inmunoterapia empírica; es necesario mantener una alta sospecha de esta entidad.

INTRODUCTION. Autoimmune encephalitis is a severe inflammatory disorder of the central nervous system caused by antibodies. Its diagnosis in pediatrics is complex, so it is relevant to deeply analyze its clinical symptoms, diagnostic methods, and treatment. OBJECTIVE. To establish the clinical symptoms, diagnosis, and treatment of Autoimmune Encephalitis in patients under 18 years of age. MATERIALS AND METHODS. Systematic review. A search for articles published in English and Spanish (2016-2021) was performed through PubMed, Google Scholar, The Lancet databases, with the following keywords: autoimmune encephalitis AND pediatrics OR children OR diagnosis AND antibodies OR treatment OR immunotherapy. The study quality was assessed using the NIH Quality Assessment Tools scale. The heterogeneity of the data only allowed the calculation of the symptom frequencies, and a narrative description of the findings was made. RESULTS. Out of 100 articles, 15 were selected. Encephalitis due to antibodies against the N-methyl-D-aspartate receptor is the most frequent. Seizures (52.6%), movement disorders (45%), and personality or behavior changes (44.4%) form the most frequent clinical symptomatology of this condition in children. The cerebrospinal fluid study showed lymphocytic pleocytosis and increased protein. Treatment with empirical immunotherapy after excluding infectious or metabolic causes is safe, compared to any other treatment. CONCLUSION. Autoimmune encephalitis against the N-methyl-D-aspartate receptor is more common in children. It has a diverse clinical presentation and improves with empirical immunotherapy; it is necessary to maintain a strong presumption of this condition.

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Neuropsychiatric syndromes in systemic lupus erythematosus (SLE) are one of the many clinical manifestations in which this pathology presents. They have a wide range of prevalence, from 37- 95% due to factors like absence of standardized definitions and non-nespecific clinical manifestations. Physiopathology is mediated by autoimmune mechanisms commonly differentiated in ischemic and inflammatory; there is a clear relationship between the pathologic pathway and the neuropsychiatric manifestation. Moreover, the blood-brain barrier plays a key role, since an alteration of the permeability allows the pass of autoantibodies to the cerebrospinal fluid. There are 19 neuropsychiatric syndromes described which include both diffuse and focal manifestations. The diagnosis must be of exclusion in sights of the more prevalent, severe and potentially deadly etiologies of the neuropsychiatric manifestations, being indispensable to conduct a full study of the patient. The therapyfocuses on symptomatic treatment for each manifestation. Immunotherapy and antithrombotic treatments should be prescripted depending on the underlying pathophysiological mechanism; however, to uncover the predominant pathological route remains a challenge. Future studies should be focused in a better understanding of the physiopathological routes in order to develop standardized diagnosis criteria and optimize an early treatment. This would have a major impact in the life of patients suffering from neuropsychiatric manifestations of SLE, whose late diagnosis is linked with greater organic damage and a poorer quality of life.

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OBJECTIVE@#To compare the consistency of programmed cell death 1-ligand 1 (PD-L1, clone E1L3N, 22C3, SP263) in different immunohistochemical staining methods.@*METHODS@#The first step was to select the optimal process: The PD-L1(clone E1L3N) antibody recommended process, self-built process ①, self-built process ② and self-built process ③ were used to perform immunohistochemical staining in 5 cases of tonsil tissue. The quality of all slides was scored by expert pathologists (0-6 points). The process with the highest score was selected. The second step was to compare the consistency between the optimal procedure and the two standard procedures. Thirty-two cases of lung non-small cell carcinoma diagnosed by pathology in Peking University First Hospital in the past two years were randomly selected. The 32 cases were stained in parallel with the SP263 and 22C3 standard procedures, and all stained slides were scored by specialized pathologists for tumor proportion score (TPS). The scoring results were grouped according to < 1%, ≥1% to < 10%, ≥10% to < 50%, and ≥50%. The consistency of PD-L1 detection antibody clone E1L3N and 22C3, E1L3N and SP263 staining results was analyzed.@*RESULTS@#Tonsil stained slides scores (0-6 points) were as follows: The recommended protocol was 5, 5, 5, 5 and 5. The self-built process ① was 5, 6, 6, 5 and 6. The self-built process ② was 4, 4, 4, 4 and 4.The self-built process ③ was 3, 3, 3, 3 and 3. The self-built process ① was the best with the highest score. The TPSs of 32 non small cell lung carcinoma (NSCLC) cases were as follows: Of self-built process ①, 6 cases were lower than 1%, 5 cases were from 1% to 10%, 10 cases were from 10% to 50%, and 11 cases were higher than 50%; of 22C3 standard procedure, 5 cases were lower than 1%, 3 cases were from 1% to 10%, 13 cases were from 10% to 50%, 11 cases were higher than 50%; of SP263 standard procedure, 7 cases were lower than 1%, 4 cases were from 1% to 10%, 11 cases were from 10% to 50%, 10 cases were higher than 50%. The results of the consistency test were as follows: The κ value for self-built process ① and 22C3 standard procedure was 0.736 (P < 0.001), the agreement was good; the κ value for self-built process ① and SP263 standard procedure was 0.914 (P < 0.001), the agreement was very good.@*CONCLUSION@#The immunostaining using PD-L1(E1L3N) with validated self-built staining protocol ① by Ventana Benchmark GX platform can obtain high quality of slides, and the TPSs based on these slides are in good agreement with 22C3 and SP263 standard procedures.

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Objective To investigate antigen optimization of Shisa like protein 1 (SHISAL1) for preparing mouse anti-human SHISAL1 polyclonal antibody and to identify the specificity of the prepared antibody. Methods Bioinformatics was employed to predict the antigenic epitope region of SHISAL1 protein, and then a polypeptide composed of amino acid residues from the site of 28 to 97 of SHISAL1, termed SHISAL1-N, was selected as the antigen. The coding region of SHISAL1-N was cloned by molecular cloning technique, and then it was inserted into pET-28a to generate pET28a-SHISAL1-N recombinant plasmid. The two recombinant plasmids pET28a-SHISAL1-N and pET28a-SHISAL1 were transformed into BL21 (DE3) bacteria and induced to express by IPTG. The two proteins were purified and immunized to female Kunming mice, respectively. The specificities and sensitivities of the acquired antibodies were detected by Western blot analysis, immunoprecipitation and immunofluorescent cytochemical staining. Results pET28a-SHISAL1-N recombinant plasmid was successfully constructed, and the two fused proteins, SHISAL1 and SHISAL1-N, were induced to express. Moreover, two types of SHISAL1 mouse polyclonal antibodies, derived from SHISAL1-N and SHISAL1 antigens, were obtained. Western blot results showed that the antibody prepared from SHISAL1 antigen was less specific and sensitive compared with the antibody prepared from SHISAL1-N antigen which could specifically identify different endogenous SHISAL1 protein. Immunoprecipitation results showed that SHISAL1-N antibody could specifically pull down SHIISAL1 protein in hepatocellular carcinoma cells and immunofluorescence results demonstrated that SHISAL1-N antibody could specifically bind to SHISAL1 protein in the cytoplasm. Conclusion We have optimized the SHISAL1 antigen and prepared the mouse anti-human SHISAL1 polyclonal antibodies successfully, which can be used for Western blot analysis, immunoprecipitation and immunofluorescence cytochemical staining.

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OBJECTIVES@#To investigate the distribution characteristics of non-bacterial pathogens in community-acquired pneumonia (CAP) in children.@*METHODS@#A total of 1 788 CAP children admitted to Shenyang Children's Hospital from December 2021 to November 2022 were selected. Multiple RT-PCR and capillary electrophoresis were used to detect 10 viral pathogens and 2 atypical pathogens, and serum antibodies of Chlamydial pneumoniae (Ch) and Mycoplasma pneumoniae (MP) were detected. The distribution characteristics of different pathogens were analyzed.@*RESULTS@#Among the 1 788 CAP children, 1 295 children were pathogen-positive, with a positive rate of 72.43% (1 295/1 788), including a viral pathogen positive rate of 59.68% (1 067/1 788) and an atypical pathogen positive rate of 22.04% (394/1 788). The positive rates from high to low were MP, respiratory syncytial virus (RSV), influenza B virus (IVB), human metapneumovirus (HMPV), human rhinovirus (HRV), human parainfluenza virus (HPIV), influenza A virus (IVA), bocavirus (BoV), human adenovirus (HADV), Ch, and human coronavirus (HCOV). RSV and MP were the main pathogens in spring; MP had the highest positive rate in summer, followed by IVA; HMPV had the highest positive rate in autumn; IVB and RSV were the main pathogens in winter. The positive rate of MP in girls was higher than that in boys (P<0.05), and there were no significant differences in other pathogens between genders (P>0.05). The positivity rates of certain pathogens differed among age groups (P<0.05): the positivity rate of MP was highest in the >6 year-old group; the positivity rates of RSV and Ch were highest in the <1 year-old group; the positivity rates of HPIV and IVB were highest in the 1 to <3 year-old group. RSV, MP, HRV, and HMPV were the main pathogens in children with severe pneumonia, while MP was the primary pathogen in children with lobar pneumonia, and MP, IVB, HMPV, RSV, and HRV were the top 5 pathogens in acute bronchopneumonia.@*CONCLUSIONS@#MP, RSV, IVB, HMPV, and HRV are the main pathogens of CAP in children, and there are certain differences in the positive rates of respiratory pathogens among children of different ages, genders, and seasons.

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OBJECTIVE@#To establish the diagnostic process of low titer blood group antibody in the occurrence of adverse reactions of hemolytic transfusion.@*METHODS@#Acid elusion test, enzyme method and PEG method were used for antibody identification. Combined with the patient's clinical symptoms and relevant inspection indexes, the irregular antibodies leading to hemolysis were detected.@*RESULTS@#The patient's irregular antibody screening was positive, and it was determined that there was anti-Lea antibody in the serum. After the transfusion reaction, the low titer anti-E antibody was detected by enhanced test. The patient's Rh typing was Ccee, while the transfused red blood cells were ccEE. The new and old samples of the patient were matched with the transfused red blood cells by PEG method, and the major were incompatible. The evidence of hemolytic transfusion reaction was found.@*CONCLUSION@#Antibodies with low titer in serum are not easy to be detected, which often lead to severe hemolytic transfusion reaction.

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Objective To prepare a rabbit anti-mouse coiled-coil domain containing 189 (Ccdc189) polyclonal antibody. Methods The pET-28a-Ccdc189 prokaryotic expression plasmid was constructed and transformed into E.coli BL21. IPTG was used to induce the expression of Ccdc189 prokaryotic protein. Adult male New Zealand rabbits were immunized with purified recombinant protein to obtain rabbit anti-mouse Ccdc189 polyclonal antibody. The specificity of the polyclonal antibody was identified by Western blot analysis, indirect ELISA and immunofluorescence histochemical staining. Results The pET-28a-Ccdc189 recombinant plasmid was successfully constructed and the expression of the Ccdc189 recombinant protein was induced. ELISA revealed that the titer of the polyclonal antibody was 1:1 000 000. Western blot and immunofluorescence staining demonstrated that the Ccdc189 polyclonal antibody could specifically identify the Ccdc189 prokaryotic protein and the Ccdc189 protein in adult wild-type mouse testis. Conclusion A polyclonal antibody with high specificity against mouse Ccdc189 was successfully created.

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The concept of "ntigen"is a relative one. The narrow concept of it condenses the process of activation of adaptive immune response and re-recognition of the same antigen, revealing the protective mechanism of vaccines with great significance for research and development of vaccines. However, the narrow concept involves adaptive immune system members: B cells, T cells and their effector products, which is difficult for beginners to understand the inherent meaning. Meanwhile, antigen classification fully summarizes the immune response process, so a variety of classification approach increases the difficulty in learning. Our teaching team analyzes the difficulties of this chapter in depth, and we implements the strategy that takes antibody structure and function as the breakthrough point and simplified adaptive immune response process as the core in teaching. A mind map that includes the main contents of this chapter is made during the process, which promotes the effectiveness of classroom teaching greatly.

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Objective To express the monkeypox virus (MPXV) A23R protein in Escherichia coli and purify by Ni-NTA affinity column, and to prepare mouse antiserum against MPXV A23R. Methods The recombinant plasmid pET-28a-MPXV-A23R was constructed and transformed into Escherichia coli BL21 to induce the expression of A23R protein. After optimizing the conditions of expression, A23R protein was highly expressed. Recombinant A23R protein was purified by Ni-NTA affinity column and identified by Western blot analysis. The purified protein was used to immunize mice for preparing the A23R polyclonal antibody, and the antibody titer was detected by ELISA. Results The expression of A23R recombinant protein reached the peak under the induced conditions of 0.6 mmol/L isopropyl-β-D-thiogalactoside (IPTG), 37 DegreesCelsius and 20 hours. The purity of the protein was about 96.07% and was identified by Western blot analysis. The mice were immunized with recombinant protein, and the titer of antibody reached 1:102 400 at the 6th week after immunization. Conclusion MPXV A23R is expressed highly and purified with a high purity and its antiserum from mouse is obtained with a high titre.

ABSTRACT

Yeast surface display (YSD) is a technology that fuses the exogenous target protein gene sequence with a specific vector gene sequence, followed by introduction into yeast cells. Subsequently, the target protein is expressed and localized on the yeast cell surface by using the intracellular protein transport mechanism of yeast cells, whereas the most widely used YSD system is the α-agglutinin expression system. Yeast cells possess the eukaryotic post-translational modification mechanism, which helps the target protein fold correctly. This mechanism could be used to display various eukaryotic proteins, including antibodies, receptors, enzymes, and antigenic peptides. YSD has become a powerful protein engineering tool in biotechnology and biomedicine, and has been used to improve a broad range of protein properties including affinity, specificity, enzymatic function, and stability. This review summarized recent advances in the application of YSD technology from the aspects of library construction and screening, antibody engineering, protein engineering, enzyme engineering and vaccine development.