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OBJECTIVE@#To investigate paraffin section thickness in immunohistochemical detection of p16 expression in cervical tissue samples.@*METHODS@#p16 immunohistochemical staining was performed in 150 cases of chronic cervicitis, 126 cases of low-grade squamous intraepithelial lesions(LSIL), 96 cases of high-grade squamous intraepithelial lesions (HSIL) and 78 cases of cervical cancer from January 2014 to March 2018 in Zhongshan Boai Hospital. The results of p16 protein expression in paraffin sections with thickness of 2, 3, 4, 5 and 6 μm were compared using Logistic regression analysis.@*RESULTS@#With the increase of slice thickness, the staining of p16 protein in nucleus gradually increased. The thickness of cervical slices in chronic cervicitis and cervical cancer samples had no significant effect on the positive rate of p16 protein(=7.817 and 1.332, both >0.05), while the thickness of slices in LSIL and HSIL samples had significant effect on the positive rate of p16 protein (=17.688 and 10.182, <0.05 or <0.01). The stable and reliable results were obtained when the slices were between 3 and 5 μm thick.@*CONCLUSIONS@#Paraffin sections with thickness of 3.0-5.0 μm are recommended for immnohistochemical staining of p16 protein in cervical tissue samples.
Subject(s)
Female , Humans , Biomarkers, Tumor , Genetics , Metabolism , Uterine Cervical Dysplasia , Cyclin-Dependent Kinase Inhibitor p16 , Genetics , Metabolism , Histocytological Preparation Techniques , Reference Standards , Immunohistochemistry , Paraffin , Squamous Intraepithelial Lesions of the Cervix , Uterine Cervical NeoplasmsABSTRACT
Objective To compare two different methods to detect the differences of gene mutation rate, sensitivity, specificity and coincidence rate of epidermal growth factor receptor (EGFR) in non-small-cell lung cancer (NSCLC) so as to assess the clinical value of qRT-PCR method and its environmental-friendly technologyplatforms.One uses environmental fixative poly hydroxyl acrylic acid and green transparent liquid dewaxing Van-clear alone or in combination to replace the traditional fixative 4％ (volume fraction) neutral buffered formalin and the traditional transparent dewaxing liquid xylene in application of quantitative real-time polymerase chain reaction (qRT-PCR).The other uses traditional reagents in direct sequencing.Methods We selected 91 cases of primary NSCLC specimens resected between May 2013 and March 2016 in Zhongshan Bo`ai Hospital and Zhongshan Hospital of Traditional Chinese Medicine.Five samples were taken from the same tumor lesion.We used a random number table to randomly divide these samples into Groups A, B , C, D, and E.Group A received direct sequencing method in detection of EGFR gene mutations.Besides, during the experiment, 4％ neutral buffered formalin was used for fixing, and xylene transparent dewaxing was used to make slices for DNA extraction dewaxing.Group B received qRT-PCR method to detect EGFR gene mutations.Meanwhile, during the experiment, 4％ neutral buffered formalin was used for fixing, and xylene transparent dewaxing was used to make slices for DNA extraction dewaxing.Group C received qRT-PCR method in detection of EGFR gene mutations.At the same time, during the experiment, polyhydroxy acrylic acid was used for fixing, and xylene transparent dewaxing was used to make slices for DNA extraction dewaxing.Group D received qRT-PCR method to detect EGFR gene mutations.In the meantime, 4％ neutral buffered formalin was used for fixing, Van-clear transparent dewaxing was used to make slices for DNA extraction dewaxing.Group E received qRT-PCR method in detection of EGFR gene mutations.In addition, during the experiment, polyhydroxy acrylic acid was used for fixing, and Van-clear transparent dewaxing was used to make slices for DNA extraction dewaxing.In addition, during the experiment, polyhydroxy acrylic acid was used for fixing, and Van-clear transparent dewaxing was used to make slices for DNA extraction dewaxing.The mutations of Exons 18, 19, 20, and 21 in EGFR genes were respectively determined in the five groups of NSCLC.Results ① Groups B, C, D, E and A did not significantly differ in the percentage of people with mutations or target site mutation rates of EGFR genes in NSCLC (P> 0.05).② The detection results of EGFR target site mutation in Groups B, C, D, E and A had good sensitivity, strong specificity, and high compliance rate.Conclusion The green transparent liquid dewaxing Van-clear alone or in combination to replace the traditional fixative 4％ neutral buffered formalin and the traditional transparent dewaxing liquid xylene in the application of qRT-PCR so as to detect EGFR gene mutations in NSCLC has good consistent results compared with the method that uses traditional reagents in direct sequencing.It has the significance and value in clinical application.
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<p><b>OBJECTIVE</b>To investigate the effects of the volume of 4% neutral phosphate buffered formalin fixative solution on the detection of human epidermal growth factor receptor 2 () gene by fluorescencehybridization (FISH) in primary invasive breast cancer.</p><p><b>METHODS</b>Tissue samples were collected from 109 patients with primary invasive breast cancer admitted in Zhongshan Boai Hospital from June 2014 to October 2016. The ratios of 4% phosphate buffered formalin fixative solution to sample volume samples were 3:1, 6:1, 9:1, 10:1, 15:1, 20:1 or 25:1 (groups A, B, C, D, E, F and G), respectively. Paraffin sections were made after 15 h of fixation. The amplification ofgene was detected by FISH. The gene amplification results ofwere observed and compared in different groups.</p><p><b>RESULTS</b>Fluorescence microscope showed that the tissue contour in groups A, B and C was vague, cell debris appeared, and the probe was positioned poorly; while the tissue contour was clear and complete in groups D, E, F and G and the probe was positioned accurately. The positive rate ofwas gradually increased from group A to D(=8.601,<0.01), and that remained stable at 24.77% in groups D to G. The positive rate of gene amplification in groups D, E, F and G was significantly higher than that in groups A, B and C (all<0.05).</p><p><b>CONCLUSIONS</b>When using FISH to detectgene in samples of primary breast invasive carcinoma, the volume of fixative solution should be at least 10 times of the sample volume to obtain accurate and stable results.</p>
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Objective To explore the correlation between histological chorioamnionitis (HC) and brain injury in preterm infants. Methods Three hundred and forty-seven cases of infants at the gestational age of 28-31 weeks who were admitted to the neonatology department of our hospital were analyzed retrospectively. They were divided into the HC group and the control group according to the pathological examination. Moreover, HC group was divided into FV group and non-FV group according to the pathological findings of fetal vasculitis (FV). Based on the findings of periodical ultrasonography, the incidences of periventricular leukomalacia (PVL), periventricular-intraventricular hemorrhage (PVH-IVH), and the PVL+PVH-IVH were compared among groups. Results The incidences of PVL in the HC group and the control group were 17.9% and 10.3%respectively. The incidences of PVL+PVH-IVH in the two groups were 5.5%and 1.48%respectively, and the difference between two groups was signiifcant (P0.05). In the HC group, the incidences of PVL in FV group and non-FV group were 28.1%and 9.87%respectively, and the difference between two groups was signiifcant (P0.05). The incidences of PVL+PVH-IVH in FV group and non-FV group were 7.81%and 3.70%respectively, and the difference between the two groups was not have signiifcant (P>0.05). Conclusions HC may increase the ncidences of PVL and PVL+PVH-IVH in the preterm infants, while its effect is minimal on PVH-IVH. FV could increase the incidence of brain injury in preterm infants.