Development and application of a method for determination of activity and vitality of Hansenula polymorpha by carboxyfluorescein diacetate-propidium iodide dua fluorescent staining / 中国生物制品学杂志

@#Objective To develop a method for determination of activity and viability of Hansenula polymorpha by dual fluorescent staining with carboxyfluorescein diacetate(CFDA)and propidium iodide(PI).Methods The time durations(5,10,20,40,60 and 120 min for CFDA,5,10,15,20 and 25 min for PI)and working solution concentrations(50,100,200and 300 μg for CFDA,2,10,20 and 30 μmol/L for PI)for the dual fluorescent staining were optimized by single factor test. Under the optimal condition,the H.polymorpha samples at theoretical survival rates of 0,25%,50%,75% and 100%were determined,of which the fluorescent intensity was observed under fluorescent microscope,and the gray value was analyzed by ImageJ software. The live and dead cells were counted,based on which the actual survival and death rates were calculated. Meanwhile,the relationships of actual gray value,actual survival rate and actual death rate to the corresponding theoretical values were analyzed. Activity and viability of three batches of cultured H.polymorpha were detected by CFDA-PI dual fluorescence staining.Results The optimal time durations for staining with CFDA and PI were 60 and 5 min,while the optimal working solution concentrations were 200 and 2 μmol/L,respectively. The actual gray value,actual survival rate and actual death rate of H.polymorpha samples at various theoretical survival rates were significantly correlated to the corresponding theoretical values(R~2=0. 998 3～0. 999 2,P < 0. 05). The CVs of activity and viability values in three detections of three batches of H.polymorpha culture were 3. 20%～4. 03% and 1. 10%～2. 27%, respectively.Conclusion The CFDA-PI dual fluorescent staining was successfully developed,which may be used for determination of activity and vitality of H.polymorpha.

Influence of blocking the short wave visible light on the stability of tear film in healthy people / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To investigate the effect of blocking short wave visible light(400-500nm)on tear film stability in healthy people. <p>METHODS: In this prospective intervention experiment, 26 college students were selected, including 12 males and 14 females, aged 22.1±1.4 years, from February to March 2020. At the same time of two adjacent days, the subjects watched the video for 1h under the same environmental conditions. For the first time, there was no short wave visible light blocking, and for the second time, they wore short wave visible light blocking myopia or flat glasses. The subjects were examined the radius of curvature of lacrimal River curved surface, tear osmolality, tear film rupture time, corneal fluorescein sodium staining score, the basic state before watching the video and after watching the video respectively. And the questionnaire of visual perception was completed at the end of the experiment. <p>RESULTS: There were significant differences in the radius of curvature of lacrimal River curved surface, tear osmolality, tear film rupture time, corneal fluorescein sodium staining score between the basic state and the short wave visible unstopped state(<i>t</i>=4.50, 5.72, 4.437, 3.245, <i>P</i><0.05). There was no significant difference between the short wave visible light blocking and the basic state(<i>t</i>=1.972, 1.993, 1.921, 1.641, <i>P</i>>0.05). There was also significant difference between the short wave visible light blocking and the short wave visible light unblocking(<i>t</i>=2.825, 3.771, 2.610, 3.028, <i>P</i>< 0.05). The results of the questionnaire showed that: eighteen subjects(69%)said that wearing the short wave visible light blocking glasses or without wearing short wave visible light blocking glasses had slight eye dryness, pain, foreign body sensation, visual fatigue and other discomfort after watching the video. Sixteen subjects(62%)said that they were more comfortable after blocking the short wave visible light. Twenty-four subjects(92%)said that they were willing to block the short wave visible light. <p>CONCLUSION: The stability of tear film could be reduced by watching the display screen for a short time. Blocking the short wave visible light could protect the stability of tear film of the ocular surface.

Clinical Observation of Pranoprofen Combined with Sodium Hyaluronate in the Treatment of Moderate and Severe Dry Eyes / 中国药房

OBJECTIVE:To explore the effectiveness and safety of pranoprofen combined with sodium hyaluronate in the treat-ment of moderate and severe dry eyes. METHODS:180 patients with moderate and severe dry eyes were divided into observation group and control group by random number table method,with 90 cases in each group. Control group was given Sodium hyaluro-nate eye drops,one drop each time,tid,and received physical therapy as cleaning eyelid,hot compress and glandulae tarsales mas-sage. Observation group was additionally given Pranoprofen eye drops,one drop each time,tid. A treatment course lasted for 2 weeks. The monocular corneal fluorescence staining score,break-up time of tear film(BUT),dry eye symptom scores,ShirmerⅠtest (SIT) before and after treatment,clinical efficacy and ADR were compared between 2 groups. RESULTS:2,4 weeks after treatment,monocular corneal fluorescence staining scores and dry eye symptom scores of 2 groups were significantly lower than be-fore treatment;BUT and SIT were significantly longer than before;those indexes after 4 weeks of treatment were significantly bet-ter than after 2 weeks of treatment;those indexes of observation group after 2,4 weeks of treatment were significantly better than those of control group at the same time,with statistical significance (P0.05). CONCLUSIONS:For moderate and severe dry eyes,prano-profen combined with sodium hyaluronate can effectively control ocular inflammation and improve tear film stability. It shows defi-nite therapeutic efficacy and good safety.

Optimization of extracting technology of total triterpenoids in Antrodia camphorata by central composite design-response surface methodology and study on antitumor activity of total triterpenoids / 中草药

Objective: To study the extracting technology of Antrodiacamphorata total triterpenoids (ACTT) and their anti-tumor activity. Methods: To optimize the ACTT extraction process by central composite design of response surface methodology (CCD-RSM).To compare the inhibition of ACTT extract at different concentration and different time on in vitroproliferation of cultured HepG2 cell using MTT assay;To observe the HepG2 cell morphology by fluorescence microscopy and flow cytometry andto detect the cell apoptosis rate by AnnexinV apoptosis assay kit. Results:The optimized process was plus 20 times amount of 90% ethanol, ultrasonic extraction twice, 40min each time, ultrasonic power of 400W.ACTT extracting rate was 11.87%, deviation between the actual and predicted values of ACTT extraction rate was 1.56%.MTT assay showed that in 12.5-200μg/mL, ACTT extract at different concentration (12.5-200μg/mL) on the proliferation of HepG2 cells showed significant inhibition(P<0.01).Microscopic observation and direct form Hoechst33342 staining were used to observethenuclear enrichment and nuclear fragmentation and other typical features of apoptosis and apoptotic bodies;Flow cytometry showed that there was significant differenceof apoptosis rate in the administration group (P<0.01) compared with the control group. Conclusion: UsingCCD-RSM to optimize antrodia ultrasonic alcohol extraction process, theactual and predicted values are consistent with high and good predictability which is feasible.ACTT extract can significantly inhibit the proliferation and induce apoptosis on HepG2 cells in vitro.

Combination of fluorescence staining and dye dyeing for mapping sentinel lymph nodes in breast cancer patients / 中国综合临床

Objective To evaluate intraoperative tracing of sentinel lymph node(SLN) by fluorescence staining combined with dye dyeing.Methods A total of 174 patients with breast cancer were randomly divided into three groups : the group A with 57 patients receiving methylene blue (MB), the group B with 58 patients receiving indocyanine green(ICG) as the lymphatic mapping tracers,the group C with 59 patients receiving MB and ICG.The sentinel and axillary lymph node of level Ⅱ, Ⅲ was excised, followed by conventional histopathology.Results There was no significant difference among the three groups in the term of visualized detection rate (x2=2.96, P =0.241).There was statistical significant difference among three groups in the term of detected lymph nodes(F=15.34, P＜0.05).Comparing with the three groups, the number of detected lymph nodes of A and B group had no significant differences(P=0.07) ,the number of detected lymph nodes of C was higher than that of A and B group, and the difference was significant(P＜0.05).There was statistical significant difference among three groups in the term of SLN positive rate (x2 =6.75, P =0.039), and there was no significant difference among A and B group(P=0.915) ,SLN positive rate of C group was higher than than A and B group, and the difference was statistical significant (P＜0.05).Conclusion Intraoperative tracing of SLN by fluorescence staining combined with dye dyeing has the skin and subcutaneous reveal advantage.The use of ICG fluorescence and MB increases lymph node detection rate.

Evaluation on effect of fluorescent staining and Ziehl-Neelsen staining methods for detecting Mycobacterium tuberculosis / 国际检验医学杂志

Objective To compare the results difference between the fluorescence staining and the acid fast staining (Ziehi‐Neelsen staining) methods in the detection of Mycobacterium tuberculosis ,and to compare the effects of the methylene blue solu‐tion ,Haris hematoxylin solution and potassium permanganate liquid as the redyeing reagents of the fluorescence staining method . Methods 198 sputum specimens collected from the patients with suspected tuberculosis symptoms and were performed the Ziehi‐Neelsen staining and the fluorescence staining respectively For comparing the difference in the detecting rate of Mycobacterium tu‐berculosis between the two kinds of method .The fluorescence staining adopted 0 .3% methylene blue solution ,0 .5% Haris hema‐toxylin solution and 0 .5% potassium permanganate solution as the redyeing reagents for comparing the effects of the fluorescence microscopic examination among different redying reagents .Results The detection rate of Mycobacterium tuberculosis was 66 .67%(132/198) for the Ziehi‐Neelsen staining ,94 .9% (188/198) for the fluorescence stainings and 94 .95% (188/198) for the methyl‐ene blue staining ,in which the detection rate of methylene blue redyeing was 94 .95% ,which of hematoxylin redyeing was 94 .44%(187/198) and which of potassium permanganate redyeing was 94 .44 (187/198) ,the differences among them were statistically sig‐nificant(P< 0 .05) .Conclusion The fluorescent staining method has the higher positive detection rate of Mycobacterium tubercu‐losis than the Ziehl‐Neelsen staining method ,in which 0 .3% methylene blue solution is a good background quenching redyeing solu‐tion .

Correlation evaluation between questionnaire and clinical examination in dry eye / 中华实验眼科杂志

Background There are two types of different questionnaires in dry eye diagnosis.But the associations about two questionnaires or questionnaire and clinical examination are still unclear.To effectively quantize the symptoms is helpful for a correct diagnosis of dry eye disease.Objective This survey was to evaluate the Standard Patient Evaluation of Eye Dryness (SPEED) and Ocular Surface Disease Index(OSDI) questionnaire for the diagnosis of dry eye and investigate the correlation between the clinical examinations and questionnaires. Methods A perspective cohort study was designed.Sixty-six patients were enrolled in this study.This study was approved by the Ethic Committee of Peking University First Hospital,and written informed consent was obtained from each subject before any ocular examination.SPEED-based and OSDI-based questionnaires were used to score the dry eye symptom and grouped according to severity of complains.Corneal fluorescence staining,tear film breakup time(BUT),Schirmer I test and tear film interferometry were performed in all patients.The correlations between two questionnaires scores and their association with clinical examinations were evaluated.Results The negative correlations were found between the SPEED-based score or OSDI-based score with BUT value(r=0.390,P=0.001 ;r=-0.395,P=0.001 ),but no significant correlations were seen between the SPEED-based score or OSDI-based score with Schirmer test( r=-0.081,P=0.515; r=-0.080,P=0.525)and tear film interferometry score(r=0.158,P=0.204;r=0.219,P=0.077).The BUT was significantly prolonged in mild symptom group compared with serious group(t=2.339,P=0.022),but no significant difference was seen in Schirmer Ⅰ test and tear film interferometry scores using SPEED-based questionnaire ( t =0.404,P =0.687 ; t =- 0.947,P =0.347 ) ; while the positive fluorescence staining rate between two groups was significantly different (x2 =0.164,P =0.685 ).When using OSDI-based questionnaire,significant difference in BUT was seen among mild,moderate and serious symptom groups ( F =11.871,P =0.000 ),and BUT in mild symptom group was delayed in comparison with moderat and serious groups( P=0.000,0.000).No significant differences were found in Schirmer Ⅰ test,tear film interferometry scores and fluorescence staining rate among three groups(F=1.432,P =0.246; F =2.799,P =0.068; x2 =6.026,P =0.050).SPEED score showed a positive correlation with OSDI score ( r =0.697,P =0.000 ). Conclusions Both OSDI and SPEED are effective tools for the evaluation of symptoms of dry eye.The two types of questionnaires are consistent in symptoms evaluation.

Comparison of Apoptosis Fluorescence Staining Assay and MIT Assay in the Antitumor Drugs Sensitivity Test / 中国肿瘤生物治疗杂志

0.05) both in AFS assay and MTT assay to detect the apoptosis of the cell lines. However, there was significant difference ( P

Effects of turmeric volatile oil on the morphology of human lung adenocarcinoma A549 cells / 第三军医大学学报

Objective To investigate the effects of turmeric volatile oil extracted from curcuma on human lung adenocarcinoma A549 cell line. Methods The morphological changes of human lung adenocarcinoma A549 cells treated by different concentrations of turmeric volatile oil were observed by inverted microscopy, Giemsa staining, and Acridine orange staining. Results ① The inverted microscopy revealed that after treatment of A549 cells with different concentrations of turmeric volatile oil for 24-72 h or specified concentration of turmeric volatile oil for 1-7 d, the lung carcinoma cells were distributed sporadically, cells were scarce, and the majority of cells were in round or oval shapes, indicating that the cell proliferation was inhibited. These changes were associated with the concentration of turmeric volatile oil. ② Giemsa staining indicated that the apoptotic rates of A549 cells were 34% and 42% respectively after turmeric volatile oil treatment at the concentrations of 0.8 ?l/ml and 1.0 ?l/ml. There was significant difference in apoptotic rate between the treatment groups and the negative control group (13.5%, P

IDENTIFICATION AND CHARACTERIZATION OF CD34~+ STEM CELLS OF HEMATOPOIETIC TUMOR CELL LINES / 解剖学报

Objective To identify and characterize the CD34~+ stem cells of hematopoietic tumor cell lines. Methods Taking CD34 molecule as a marker to identify if there are CD34~+ tumor cells in hematopoietic tumor cell lines(THP-1,MEL,K562,MOLT-4,RAJI,RPMI8226,SP2/0 and DCS) by fluorescence staining and FCM technology. We separated CD34~+ cells from K562 and MOLT-4 for further study.CD34~+ cells were cultured and observed their development.CD34~+ and CD34 +- cells were compared in their functional status(mRNA),cell cycles(G -0/G -1),their degree of differentiation(Id-1 expression) and their multi-drug resistance(gp expression). Results The percentages of CD34~+ cells were 6