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Introducción: La citometría de flujo permite la cuantificación de las subpoblaciones de linfocitos con una elevada sensibilidad, especificidad y objetividad. Estas ventajas solo se logran con un proceso laborioso de diseño individualizado y controlado para cada experimento. Objetivo: Diseñar un protocolo de un solo tubo policromático de citometría flujo para inmunofenotipo linfocitario periférico. Métodos: Se realizó un estudio experimental in vitro con muestras de sangre periférica obtenidas de tres voluntarios sanos, en el Centro Nacional de Genética Médica, en marzo de 2019. El tubo se compuso de seis marcadores de linaje para identificar linfocitos B, T, natural killer y natural killer T. Se desarrolló un protocolo de lisis de hematíes sin lavado. Se emplearon anticuerpos monoclonales conjugados con fluorocromos. El punto óptimo de concentración correspondió al mayor índice de tinción y conservación de los porcentajes de positividad de cada población. Se realizó la construcción progresiva del tubo y se propuso una estrategia lógica de secuencia de ventanas para el análisis de datos. Resultados: Los marcadores seleccionados permitieron realizar correctamente el inmunofenotipo linfocitario periférico. En los cinco puntos de titulación se observaron buenas discriminaciones entre las señales positivas y negativas, excepto para el anti-CD56 que presentó una tendencia decreciente del índice de tinción. El volumen total de conjugados requeridos para la determinación de los 6 antígenos fue de 3,75 μL por tubo. Conclusiones: Se obtuvo un tubo policromático que permite el inmunofenotipo periférico de forma rápida y precisa por seis antígenos linfocitarios simultáneamente, con el empleo de pequeños volúmenes de conjugado y sangre(AU)
Introduction: Flow cytometry allows quantification of lymphocyte subpopulations with high sensitivity, specificity and objectivity. These advantages are only achieved through the hardworking process of individualized and controlled design for each experiment. Objective: To design a flow cytometry protocol of a single polychromatic tube for peripheral lymphocyte immunophenotype. Methods: An experimental in vitro study was carried out, in March 2019, with peripheral blood samples obtained from three healthy volunteers, at the National Center for Medical Genetics. The tube was made up of six lineage markers for identifying natural B and T lymphocytes, natural killers and natural killer T cells. A protocol was developed for red blood cell lysis without washing. Fluorochrome-conjugated monoclonal antibodies were used. The optimal point of concentration corresponded to the highest staining index and preservation of the positivity percentages of each population. Progressive tube construction was performed and a logical window sequence strategy was proposed for data analysis. Results: The chosen markers allowed to carry out correct peripheral lymphocyte immunophenotype. Good discriminations between positive and negative signals were observed at the five titration points, except for anti-CD56, which presented a decreasing trend in the staining index. The total volume of conjugates required for determination of the six antigens was 3.75 μL per tube. Conclusions: A polychromatic tube was obtained that allows to carry out peripheral immunophenotype quickly and precisely by six lymphocyte antigens simultaneously, with the use of small volumes of conjugate and blood(AU)
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Humans , Process Optimization , Flow Cytometry/methods , Genetics, Medical , Construction IndustryABSTRACT
This study was aimed to discuss the anti-inflammatory and immunization properties ofRe-Du-Ning (RDN) injection in the treatment of acute exacerbation of chronic obstructive pulmonary disease (AECOPD) with phlegm-heat stagnated in the lung syndrome. A total of 110 in-patients were collected from January 2012 to December 2013. Cases were randomly divided into the treatment group (basic treatment plan + 20 ml RDN injection + 250 mL of 5% GS injection, once a day, intravenous injection) and the control group (basic treatment plan + 20 ml of 0.9% NS injection + 250 mL of 5% GS injection, once a day, intravenous injection), with 55 cases in each group. The treatment course was 14 days. Observations were made on traditional Chinese medicine (TCM) symptom score, clinical effect, blood routine examination, blood gas analysis and T lymphocyte subgroups before and after the treatment in both groups. The results showed that in the aspects of TCM symptom score and clinical effect, the posttreatment TCM symptom score and total integration in the treatment group were obviously improved compared to the control group except for shortness of breath, dry mouth and thirsty, flushing (P < 0.05). The total effective rate was obviously better than that of the control group. In the aspect of anti-inflammation, the total white blood cell (WBC) count and the normal neutrophil percentage of both pretreatment and posttreatment in the treatment group were obviously decreased compared to pretreatment (P < 0.05). However, there were no statistical differences on WBC count and normal neutrophil percentage compared to posttreatment in the control group. In the aspect of blood gas analysis, the posttreatment PaCO2 and PaO2 of the treatment group were obviously better than the control group (P < 0.05). In the aspect of immune regulation, the posttreatment T lymphocyte subgroups CD3+ and CD4+ of the treatment group were higher than the control group. The expression inhibition / cytotoxic lymphocyte (CD8+) was obviously lower than the control group (P < 0.05). The CD4+ / CD8+ was back to the normal reference level. It was concluded that in the treatment of AECOPD with phlegm-heat stagnated in the lung syndrome, on the basis of western medicine symptomatic treatment plan, RDN injection was assisted to clear heat, relieve toxin and remove phlegm. It can obviously improve patient’s clinical symptoms and increase the clinical therapeutic effects. The treatment was especially targeted to infection-induced respiratory failure patients combined low immunity with possible identified therapeutic effects.
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Objective To discuss the metabolic characteristics and significance of peripheral blood cell and T lymphocyte subgroup in patients with air condition disease.Methods Thirty-six patients diagnosed as air condition diseases with fever and 30 patients diagnosed as non-fever patients enrolled this study as observation group(fever group and non-fever group respectively),and 30 health population were used as control.The changes of peripheral blood cell and T lymphocyte was measured using regular blood examination and flow cytometry.Results(1)No significant diffence on white blood cell was found between the three groups(F =1.89,P =0.134);(2)The percentage of neutrophils was(68.13 ±5.67)％,(53.24 ±8.36)％ and(61.25 ±6.31)％ respectively in the fever,non-fever and control group,respectively.Compared to the non-fever and control group,the percentage of neutrophils were significantly higher(t =6.64,9.13; P =0.018,0.027 respectively).No signiciant difference in the percentage of neutrophils was found between the non-fever group and control group(P =0.356,0.125).The percentage of lymphocyte percentage was(31.66 ±7.11)％,in the fever group,which was significantly higher than those of(43.67 ±8.17)％ and(30.98 ± 8.76)％ in the nonfever and control group(t =5.59,4.98 ; P =0.022,0.021 respectively).(3)The percentage of CD4 + was (36.351 ± 6.531)％,(42.676 ± 5.169)％ and(39.253 ± 5.461)％ in the fever,non-fever and control group,respectively.The percentage of CD4 + in the fever group was significantly lower than that in the non-fever group(t =1.28,P =0.039),whereas the percentage of CD4 + in the non-fever group was significantly higher than that in the control group(t =3.80,P =0.048).The percentage of CD8 + was(28.084 ± 7.344)％,(21.186 ± 6.331)％ and(24.607 ± 7.108)％,respectively.Compared to nonfever group,the percentage of CD8+ was significantly higher(t =3.17,P =0.016).(4)The ratio of CD4 +/CD8 + was(1.629 ±0.554)％、(2.380 ± 0.391)％,and(2.079 ± 0.659)％.Compared to the non-fever and control group,the ration in the fever group was siginificantly lower(t =19.21,P ＜ 0.001 ; t =6.98 ; P =0.006),whereas compared to the control group,the ratio in the non-fever group was significantly higher(t =8.68,P =0.031).Conclusion The changes of T subgroup show the unbalance immunity function network in patients with air condition disease.
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Objective: To study the mechanism of Ainsliaea fragrans Champ in treating cervicitis.Methods: Latex rubber of oxybenzene was adopted to establish the rats' model of cervicitis by endovaginal injection.After 4 and 10 days,the effect of Ainsliaea fragrans Champ on PGE2 expression in cervix mucosae and the subgroups of T lymphocyte in peripheral blood of rats was observed,and the histopathologic examination was done.Results: omparing with the model group,Ainsliaea fragrans Champ can significantly decrease the PGE2 levels in rats' cervix mucosae after 4 days(P
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Objective:To gain an insight into the immunological mechanism of tuberculosis by observing the variation of T lymphocyte subgroups in endobronchial tuberculosis(EBTB) patients.Methods:The variation of T lymphocyte subgroups was determined by flow cytometry in 23 EBTB patients and 25 physical examinees and results were compared.Results:The value of CD8 T lymphocytes was significantly lower in the EBTB patients than in healthy controls(P0.05).Conclusion:The pathogenesis of tuberculosis is associated with the changes in the immune system.CD4 and CD8 T lymphocytes play an important role in the cell-mediated immune response.