Journal: Biotechniques

Article Title: Generation of genetically tailored porcine liver cancer cells by CRISPR/Cas9 editing

doi: 10.2144/btn-2020-0119

Figure Lengend Snippet: Oligonucleotides used in the study.

Article Snippet: The following anti-ARID1A primary antibodies were used: #sc-373784 and #sc-32761 (Santa Cruz Biotechnolog, TX, USA) and #A301-041A (Bethyl Laboratories, TX, USA).

Techniques: Sequencing

Journal: Biotechniques

Article Title: Generation of genetically tailored porcine liver cancer cells by CRISPR/Cas9 editing

doi: 10.2144/btn-2020-0119

Figure Lengend Snippet: (A) Schematic representation of the porcine ARID1A locus, showing the location of spacer sequences crRNA#1 and crRNA#2 (underlined blue font). Protospacer-adjacent motif sequences are marked in red. The introns are represented by lines, coding regions of exons are shaded in black and the untranslated region is white. (B) Comparing the CRISPR/Cas9-mediated editing efficiency of two ARID1A -targeting gRNAs. Porcine HCC line-1 cells were transfected with 15 nM RNP comprising Cas9 and gRNA#1 or gRNA#2. Nontransfected cells were used as control. Genomic DNA was collected 2 days post-transfection and analyzed by targeted NGS. The bar graph depicts the percentages (%) of total reads that displayed indels at the gRNA target site occurring as a result of nonhomologous end joining. (C & D) Top 15 reads detected by targeted NGS analysis, mapped to the reference sequence. The percentages of reads for each sequence are shown on the right. The asterisk indicates non-edited reads. Dashed line: predicted Cas9 cleavage position; red box: insertion; dash: deleted base. HCC: Hepatocellular carcinoma; NGS: Next-generation sequencing; RNP: Ribonucleoprotein.

Article Snippet: The following anti-ARID1A primary antibodies were used: #sc-373784 and #sc-32761 (Santa Cruz Biotechnolog, TX, USA) and #A301-041A (Bethyl Laboratories, TX, USA).

Techniques: CRISPR, Transfection, Control, Sequencing, Next-Generation Sequencing

Journal: Biotechniques

Article Title: Generation of genetically tailored porcine liver cancer cells by CRISPR/Cas9 editing

doi: 10.2144/btn-2020-0119

Figure Lengend Snippet: (A) Schematic representation of the workflow to sort RNP-transfected cells. A tracrRNA conjugated with ATTO-550 fluorescent dye was mixed with crRNA to form gRNA. The fluorescently labeled gRNA and purified Cas9 were complexed to form a ribonucleoprotein complex (RNP) which was then transfected into cells, allowing visual assessment of transfection efficiency using a fluorescence microscope. 48 h later, the cells were sorted by FACS to separate successfully transfected fluorescent cells. (B) ARID1A editing efficiency in unsorted or FACS-sorted porcine HCC cells. Porcine HCC line-1 cells were transfected with 15 nM RNP comprising Cas9 and ARID1A targeting gRNA#1. Nontransfected control cells were also analyzed. FACS-sorted cells were used for genomic analysis directly after sorting. The bar graph depicts the percentages (%) of total reads that displayed indels at the gRNA target site as analyzed by NGS. HCC: Hepatocellular carcinoma; NGS: Next-generation sequencing.

Article Snippet: The following anti-ARID1A primary antibodies were used: #sc-373784 and #sc-32761 (Santa Cruz Biotechnolog, TX, USA) and #A301-041A (Bethyl Laboratories, TX, USA).

Techniques: Transfection, Labeling, Purification, Fluorescence, Microscopy, Control, Next-Generation Sequencing

Journal: Biotechniques

Article Title: Generation of genetically tailored porcine liver cancer cells by CRISPR/Cas9 editing

doi: 10.2144/btn-2020-0119

Figure Lengend Snippet: (A) ARID1A editing efficiency in four porcine HCC cell lines following transfection with 25 nM ribonucleoprotein comprising Cas9 and gRNA#1. Genomic DNA was collected 2 days post-transfection and analyzed by targeted NGS. The bar graph depicts the percentages (%) of total reads that displayed indels at the gRNA target site occurring as a result of nonhomologous end joining. (B) Top 15 reads detected by targeted NGS analysis in the four porcine HCC lines, mapped to the reference sequence. The identity and frequency of indels in the four cell lines is consistent. The percentages of reads of each sequence are shown on the right. The location of the indel relative to the expected cleavage site is shown on the left, followed by a colon and the number of nucleotides inserted (i) or deleted (d). The asterisk denotes non-edited reads. Dashed line: predicted Cas9 cleavage position; red box: insertion; dash: deleted base. HCC: Hepatocellular carcinoma; NGS: Next-generation sequencing.

Article Snippet: The following anti-ARID1A primary antibodies were used: #sc-373784 and #sc-32761 (Santa Cruz Biotechnolog, TX, USA) and #A301-041A (Bethyl Laboratories, TX, USA).

Techniques: Transfection, Sequencing, Next-Generation Sequencing

Journal: Biotechniques

Article Title: Generation of genetically tailored porcine liver cancer cells by CRISPR/Cas9 editing

doi: 10.2144/btn-2020-0119

Figure Lengend Snippet: (A) Sanger sequencing chromatograms of porcine HCC cells transfected with Cas9 and ARID1A gRNA#1 and control nontransfected cells. Overlapping peaks near the gRNA target site are observed for the sample subjected to CRISPR/Cas9 editing. Dashed line: predicted Cas9 cleavage position; underlined nucleotides: crRNA spacer sequence. (B) Top 5 reads detected by ICE software analysis of Sanger sequencing data in three porcine HCC lines transfected with Cas9 and ARID1A gRNA#1. The type and frequency of indels is shown on the left and is consistent among the three cell lines. (C) The editing efficiency (%) and frequency of the top five frequent reads (%) identified by ICE software analysis of Sanger sequencing data as compared with NGS analysis in three ARID1A -edited porcine HCC cells. The location of the indel relative to the expected cleavage site is shown on the left, followed by a colon and the number of nucleotides inserted (i) or deleted (d). Overall similarity was noted between the two analysis methods. HCC: Hepatocellular carcinoma; NGS: Next-generation sequencing.

Article Snippet: The following anti-ARID1A primary antibodies were used: #sc-373784 and #sc-32761 (Santa Cruz Biotechnolog, TX, USA) and #A301-041A (Bethyl Laboratories, TX, USA).

Techniques: Sequencing, Transfection, Control, CRISPR, Software, Next-Generation Sequencing

Journal: Biotechniques

Article Title: Generation of genetically tailored porcine liver cancer cells by CRISPR/Cas9 editing

doi: 10.2144/btn-2020-0119

Figure Lengend Snippet: (A) Possible editing outcomes of CRISPR/Cas9 include the following: an identical indel is introduced in both alleles (homozygous mutation), different indels occur in each allele (biallelic mutation) or an indel is observed in an allele while the other allele remains unedited (heterozygous mutation). (B–D) Analysis of representative single cells clones isolated from porcine HCC cells transfected with Cas9 and ARID1A gRNA#1. (B) Reads detected by targeted NGS analysis mapped to the reference sequence (top) for three single cell clones. Dashed line: predicted Cas9 cleavage position; red box: insertion; dash: deleted base. (C) The predicted translation of ARID1A protein for the representative clones. The dotted region represents amino acids with frameshift mutation. (D) Arginase-1 staining (green) merged with DAPI staining (blue) for parental porcine HCC cell line and the representative single-cell clones (scale bar = 200 μm). All clones show positive arginase-1 staining, confirming their hepatocellular origin. HCC: Hepatocellular carcinoma; NGS: Next-generation sequencing.

Article Snippet: The following anti-ARID1A primary antibodies were used: #sc-373784 and #sc-32761 (Santa Cruz Biotechnolog, TX, USA) and #A301-041A (Bethyl Laboratories, TX, USA).

Techniques: CRISPR, Mutagenesis, Clone Assay, Isolation, Transfection, Sequencing, Staining, Next-Generation Sequencing

Journal: Biotechniques

Article Title: Generation of genetically tailored porcine liver cancer cells by CRISPR/Cas9 editing

doi: 10.2144/btn-2020-0119

Figure Lengend Snippet: Mutations near the expected Cas9 cleavage site of ARID1A in single-cell clones isolated from porcine hepatocellular carcinoma cells treated with Cas9 and gRNA#1 targeting ARID1A .

Article Snippet: The following anti-ARID1A primary antibodies were used: #sc-373784 and #sc-32761 (Santa Cruz Biotechnolog, TX, USA) and #A301-041A (Bethyl Laboratories, TX, USA).

Techniques: Clone Assay, Isolation, Knock-Out, Mutagenesis

Journal: Oncogene

Article Title: Next-generation sequencing of endoscopic biopsies identifies ARID1A as a tumor-suppressor gene in Barrett’s esophagus

doi: 10.1038/onc.2012.586

Figure Lengend Snippet: Tier 1 single-nucleotide variants

Article Snippet: 59 Primary antibodies against ARID1A (A301–041A, Bethyl Laboratories, Montgomery, TX, USA, dilution 1:500) and α-tubulin (protein loading control, Santa Cruz, dilution 1:30 000) were used.

Techniques: Variant Assay

Journal: Oncogene

Article Title: Next-generation sequencing of endoscopic biopsies identifies ARID1A as a tumor-suppressor gene in Barrett’s esophagus

doi: 10.1038/onc.2012.586

Figure Lengend Snippet: Immunohistochemistry for ARID1A (left panels) and p53 (right panels) in representative cores of BE (a, b, 20 × ), EAC (c, d, 20 × ) and lymph node metastasis (LNM; e, f, 20 ×) lesions. In all lesions, nuclear ARID1A expression is completely lost, whereas there is nuclear accumulation of p53. The NSE in image (c, d, black arrows) served as control for staining as ARID1A is present and p53 is absent. The red arrows (c, d) indicate EAC.

Article Snippet: 59 Primary antibodies against ARID1A (A301–041A, Bethyl Laboratories, Montgomery, TX, USA, dilution 1:500) and α-tubulin (protein loading control, Santa Cruz, dilution 1:30 000) were used.

Techniques: Immunohistochemistry, Expressing, Control, Staining

Journal: Oncogene

Article Title: Next-generation sequencing of endoscopic biopsies identifies ARID1A as a tumor-suppressor gene in Barrett’s esophagus

doi: 10.1038/onc.2012.586

Figure Lengend Snippet: (a) Immunohistochemistry results for lesions in which complete ARID1A loss was detected. Nuclear accumulation of p53 was observed in 5/12 EAC ARID1A loss patients. ARID1A loss was observed in 0% (0/76), 4.9% (2/41), 14.3% (4/28), 16% (8/50), 12.2% (12/98) and 6.5% (2/31) of the NSE, BE, LGD, HGD, EAC and lymph node metastasis (LNM), respectively (b). There was a stepwise increase in presence of nuclear accumulation of p53 seen during neoplastic progression of BE (c).

Article Snippet: 59 Primary antibodies against ARID1A (A301–041A, Bethyl Laboratories, Montgomery, TX, USA, dilution 1:500) and α-tubulin (protein loading control, Santa Cruz, dilution 1:30 000) were used.

Techniques: Immunohistochemistry

Journal: Oncogene

Article Title: Next-generation sequencing of endoscopic biopsies identifies ARID1A as a tumor-suppressor gene in Barrett’s esophagus

doi: 10.1038/onc.2012.586

Figure Lengend Snippet: Robust ARID1A expression was detected at the expected size in OE33, JHesoAD1 and HEEpiC cells. ARID1A protein was efficiently knocked down (81% compared with mock OE33) in OE33 cells (a). A significant increase in cell growth (b) and promotion of invasion (c) was observed in the ARID1A KD OE33 cells. The brown staining in photo (d, 20 × ) and (e, 20 × ), which are example photos, depicts incorporation of BrdU in mock and ARID1A KD cells, respectively. Overall, 48.8% and 67.9% of the mock and ARID1A KD cells, respectively, were proliferative 36–48 h after transfection (P <0.001). (f) Shows quantitative reverse transcriptase (qRT)-PCR validation results (four biological replicates) of the microarray experiment. The error bar in the graph indicates the standard error of the mean, whereas the asterisk depicts a P <0.05 between ARID1A KD and mock OE33 cells.

Article Snippet: 59 Primary antibodies against ARID1A (A301–041A, Bethyl Laboratories, Montgomery, TX, USA, dilution 1:500) and α-tubulin (protein loading control, Santa Cruz, dilution 1:30 000) were used.

Techniques: Expressing, Staining, Transfection, Reverse Transcription, Quantitative RT-PCR, Biomarker Discovery, Microarray

Journal: Oncogene

Article Title: Next-generation sequencing of endoscopic biopsies identifies ARID1A as a tumor-suppressor gene in Barrett’s esophagus

doi: 10.1038/onc.2012.586

Figure Lengend Snippet: ARID1A mutations in solid carcinomas

Article Snippet: 59 Primary antibodies against ARID1A (A301–041A, Bethyl Laboratories, Montgomery, TX, USA, dilution 1:500) and α-tubulin (protein loading control, Santa Cruz, dilution 1:30 000) were used.

Techniques:

Journal: Oncogene

Article Title: Next-generation sequencing of endoscopic biopsies identifies ARID1A as a tumor-suppressor gene in Barrett’s esophagus

doi: 10.1038/onc.2012.586

Figure Lengend Snippet: Summary frequent mutations in EAC discovered by NGS

Article Snippet: 59 Primary antibodies against ARID1A (A301–041A, Bethyl Laboratories, Montgomery, TX, USA, dilution 1:500) and α-tubulin (protein loading control, Santa Cruz, dilution 1:30 000) were used.

Techniques: Mutagenesis

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: Univariate correlation analysis of ARID1A with clinicopathological variables.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques: Mutagenesis

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: Representative immunohistochemistry images of important markers. (A) Diffuse strong expression of ARID1A; (B) reduced expression of ARID1A; (C) heterogeneous expression of ARID1A; (D) complete loss expression of ARID1A; (E) positive expression of MLH1; (F) negative expression of MLH1; (G) positive expression of E‐cadherin; (H) negative expression of E‐cadherin; (I) wild‐type expression of p53 (nuclear staining of variable intensity); (J) mutant‐type expression of p53 (diffuse and uniform strong nuclear staining); (K) mutant‐type expression of p53 (complete absence of nuclear staining); (L) positive expression of EBV. EBER, EBV‐encoded RNA; EBER‐ISH, EBV‐encoded RNA in situ hybridization.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques: Immunohistochemistry, Expressing, Staining, Mutagenesis, RNA In Situ Hybridization

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: (A) Flowchart of TCGA molecular classification based on IHC and EBER‐ISH. (B) Distribution of TCGA subtypes. (C) ARID1A expression in TCGA subtypes. CIN, chromosomal instability; EBV, Epstein–Barr virus; GS, genomically stable; MSI, microsatellite instability; ns, no significance. ***: p < 0.001.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques: Expressing, Virus

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: Correlation analysis of ARID1A. (A) Variables screened by multivariate logistic regression analysis. (B) and (C) Variables filtered out by the minimalist model of LASSO regression. (D) Variables ranked by importance using random forest. dMMR, mismatch repair deficient; EBER, EBV‐encoded RNA; MMR: mismatch repair; PD‐L1, programmed cell death ligand‐1; pMMR, mismatch repair proficient.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques:

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: Survival analysis plots. (A) Survival analysis by ARID1A expression in the entire cohort. (B) Survival analysis by TCGA subtypes in the entire cohort. (C)–(F) Survival analysis by ARID1A expression in TCGA subtypes. CIN, chromosomal instability; EBV, Epstein–Barr virus; GS, genomically stable; MSI, microsatellite instability. ***: p < 0.001.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques: Expressing, Virus

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: Univariate and multivariate Cox regression analysis for overall survival in GS subtype.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques: Significance Assay

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: Heatmaps of CD4, CD8, and PDL1 expression and bar plots of TCGA subtypes distribution. (A) Heatmap of CD4, CD8, and PD‐L1 expression in tumor center. (B) Distribution of TCGA subtypes in ARID1A negative and positive groups in tumor center. (C) Heatmap of CD4, CD8, and PD‐L1 expression in invasive margin. (D) Distribution of TCGA subtypes in ARID1A negative and positive groups in invasive margin. CIN, chromosomal instability; EBV, Epstein–Barr virus; GS, genomically stable; IM, invasive margin; MSI, microsatellite instability; TC, tumor center; TCGA, The Cancer Genome Atlas. ***: p < 0.001.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques: Expressing, Virus

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: Impact of ARID1A status on the expression of CD4, CD8, and PD‐L1. (A) The entire mIF cohort in tumor center; (B)–(E) TCGA subtypes in tumor center; (F) the entire mIF cohort in invasive margin; (G)–(J) TCGA subtypes in invasive margin. CIN, chromosomal instability; EBV, Epstein–Barr virus; GS, genomically stable; IM, invasive margin; MSI, microsatellite instability; ns, no significance; TC, tumor center. *: p < 0.05; **: p < 0.01; ***: p < 0.001.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques: Expressing, Virus

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: The correlation between PD‐L1 expression of IHC and immune infiltration. (A) and (D) The PD‐L1 expression in IHC was consistent with that in mIF; The expression of CD4/CD8 in the PD‐L1 positive group was significantly higher than that in PD‐L1 negative group for all cases. (B) and (E) When ARID1A was negative, the expression of CD4/CD8 in the PD‐L1 positive group was significantly higher than that in PD‐L1 negative group. (C) and (F) When ARID1A was positive, there was no difference in the expression of CD4/CD8 between PD‐L1 negative and PD‐L1 positive groups. IM, invasive margin; mIF, multiplex immunofluorescence; ns, no significance; TC, tumor center. *: p < 0.05; **: p < 0.01; ***: p < 0.001.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques: Expressing, Multiplex Assay, Immunofluorescence

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: The correlation between PD‐L1 expression, CD4 + , and CD8 + T cell infiltration in mIF. (A) and (D) PD‐L1, CD4, and CD8 were positively correlated in all cases; (B) and (E) PD‐L1, CD4, and CD8 were positively correlated in ARID1A negative cases; (C) and (F) CD4 and CD8 were positively correlated, while PD‐L1 was not associated with CD4 and CD8 in ARID1A positive cases. Corr, correlation coefficients; IM, invasive margin; ns, no significance; TC, tumor center; *: p < 0.05; **: p < 0.01; ***: p < 0.001.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques: Expressing

Journal: Cancer Medicine

Article Title: Prognostic and immune infiltration significance of ARID1A in TCGA molecular subtypes of gastric adenocarcinoma

doi: 10.1002/cam4.6294

Figure Lengend Snippet: Representative mIF images according to the expression status of ARID1A in the tumor center and invasive margin. IM ARID1A‐Neg, invasive margin and ARID1A negative; IM ARID1A‐Pos, invasive margin and ARID1A positive; TC ARID1A‐Neg, tumor center and AIRD1A negative; TC ARID1A‐Pos, tumor center and AIRD1A positive.

Article Snippet: The primary antibodies' information were as follows, ARID1A (EPR13501, Rabbit, 1:1000, Abcam), E‐cadherin (EP6, Rabbit, Prediluted, Origene), MSH2 (RED2, Rabbit, Prediluted, Origene), MSH6 (EP49, Rabbit, Prediluted, Origene), MLH1 (ES05, Rabbit, Prediluted, Origene), PMS2 (EP51, Rabbit, Prediluted, Origene), p53 (DO‐7, Mouse, Prediluted, Origene), PD‐L1 (SP263, Rabbit, Prediluted, Roche), Ki‐67 (MyM1‐Ki67, Mouse, Prediluted, Anbiping), and Her‐2 (4B5, Rabbit, Prediluted, Roche).

Techniques: Expressing

Journal: PLoS ONE

Article Title: Decreased Expression of the ARID1A Gene Is Associated with Poor Prognosis in Primary Gastric Cancer

doi: 10.1371/journal.pone.0040364

Figure Lengend Snippet: The relative mRNA expression of ARID1A was significantly decreased in gastric cancer tissues compared with the matched adjacent nontumorous tissues (n = 66, P = 0.0029). Horizontal lines represent the mean.

Article Snippet: The sections were then incubated with a mouse monoclonal antibody against ARID1A (Abgent Primary Antibody Company, USA, at a 1∶300 dilution) at 4°C overnight and then incubated with horseradish peroxidase (HRP) (ChemMateTM DAKO EnVisionTM Detection Kit) at room temperature for 30 min. After washing in PBS, the visualization signal was developed with 3, 3′-diaminobenzidine (DAB) solution, and all of the slides were counterstained with hematoxylin.

Techniques: Expressing

Journal: PLoS ONE

Article Title: Decreased Expression of the ARID1A Gene Is Associated with Poor Prognosis in Primary Gastric Cancer

doi: 10.1371/journal.pone.0040364

Figure Lengend Snippet: (A) Relative ARID1A protein expression levels in gastric cancer tissues and noncancerous tissues (ARID1A/GAPDH, n = 25, P = 0.0015). Horizontal lines represent the mean. (B) Representative result of ARID1A protein expression in 4 paired gastric tumorous and the matched adjacent nontumorous tissues (C, gastric cancer tissues; N, matched noncancerous gastric mucosa). (C) The ARID1A protein level was remarkably decreased in gastric cancer cell lines, SGC7901, AGS, especially in MGC803, compared with normal gastric cell line GES1.

Article Snippet: The sections were then incubated with a mouse monoclonal antibody against ARID1A (Abgent Primary Antibody Company, USA, at a 1∶300 dilution) at 4°C overnight and then incubated with horseradish peroxidase (HRP) (ChemMateTM DAKO EnVisionTM Detection Kit) at room temperature for 30 min. After washing in PBS, the visualization signal was developed with 3, 3′-diaminobenzidine (DAB) solution, and all of the slides were counterstained with hematoxylin.

Techniques: Expressing

Journal: PLoS ONE

Article Title: Decreased Expression of the ARID1A Gene Is Associated with Poor Prognosis in Primary Gastric Cancer

doi: 10.1371/journal.pone.0040364

Figure Lengend Snippet: (A) Strong ARID1A staining was observed in noncancerous gastric mucosa. (B) ARID1A-negative gastric adenocarcinoma, Grade 3. (C) Weak ARID1A staining in gastric adenocarcinoma, Grade 2. (D) Strong ARID1A staining in gastric adenocarcinoma, Grade 1.

Article Snippet: The sections were then incubated with a mouse monoclonal antibody against ARID1A (Abgent Primary Antibody Company, USA, at a 1∶300 dilution) at 4°C overnight and then incubated with horseradish peroxidase (HRP) (ChemMateTM DAKO EnVisionTM Detection Kit) at room temperature for 30 min. After washing in PBS, the visualization signal was developed with 3, 3′-diaminobenzidine (DAB) solution, and all of the slides were counterstained with hematoxylin.

Techniques: Staining

Journal: PLoS ONE

Article Title: Decreased Expression of the ARID1A Gene Is Associated with Poor Prognosis in Primary Gastric Cancer

doi: 10.1371/journal.pone.0040364

Figure Lengend Snippet: Correlation between ARID1A expression and clinicopathological variables of 224 gastric cancer cases.

Article Snippet: The sections were then incubated with a mouse monoclonal antibody against ARID1A (Abgent Primary Antibody Company, USA, at a 1∶300 dilution) at 4°C overnight and then incubated with horseradish peroxidase (HRP) (ChemMateTM DAKO EnVisionTM Detection Kit) at room temperature for 30 min. After washing in PBS, the visualization signal was developed with 3, 3′-diaminobenzidine (DAB) solution, and all of the slides were counterstained with hematoxylin.

Techniques: Expressing

Journal: PLoS ONE

Article Title: Decreased Expression of the ARID1A Gene Is Associated with Poor Prognosis in Primary Gastric Cancer

doi: 10.1371/journal.pone.0040364

Figure Lengend Snippet: The survival rate of patients in the ARID1A-negative group was significantly lower than that of patients in the ARID1A-positive group (log-rank test, P = 0.003).

Article Snippet: The sections were then incubated with a mouse monoclonal antibody against ARID1A (Abgent Primary Antibody Company, USA, at a 1∶300 dilution) at 4°C overnight and then incubated with horseradish peroxidase (HRP) (ChemMateTM DAKO EnVisionTM Detection Kit) at room temperature for 30 min. After washing in PBS, the visualization signal was developed with 3, 3′-diaminobenzidine (DAB) solution, and all of the slides were counterstained with hematoxylin.

Techniques:

Journal: PLoS ONE

Article Title: Decreased Expression of the ARID1A Gene Is Associated with Poor Prognosis in Primary Gastric Cancer

doi: 10.1371/journal.pone.0040364

Figure Lengend Snippet: Univariate and multivariate analyses of overall survival of gastric cancer patients.

Article Snippet: The sections were then incubated with a mouse monoclonal antibody against ARID1A (Abgent Primary Antibody Company, USA, at a 1∶300 dilution) at 4°C overnight and then incubated with horseradish peroxidase (HRP) (ChemMateTM DAKO EnVisionTM Detection Kit) at room temperature for 30 min. After washing in PBS, the visualization signal was developed with 3, 3′-diaminobenzidine (DAB) solution, and all of the slides were counterstained with hematoxylin.

Techniques:

Journal: PLoS ONE

Article Title: Decreased Expression of the ARID1A Gene Is Associated with Poor Prognosis in Primary Gastric Cancer

doi: 10.1371/journal.pone.0040364

Figure Lengend Snippet: (A) Western blotting analysis of restoring ARID1A expression in MGC803 cells. (B) Western blotting analysis of silenced ARID1A expression in GES1 cells. (C) Cell proliferation assay showing the suppressive effect of restoring ARID1A expression on the in vitro proliferation of MGC803 cell line. (D) ARID1A inhibited colony formation of MGC803 cells. Images are shown on the left and on the right, quantitative analyses of plaque numbers are shown as mean ± SD. (E) Cell proliferation assay showing significantly enhanced proliferation rate of ARID1A -silenced GES1 cells compared with mock siRNA treatment GES1 cells. *, P <0.05 versus the mock-control; **, P <0.01 versus the mock-control.

Article Snippet: The sections were then incubated with a mouse monoclonal antibody against ARID1A (Abgent Primary Antibody Company, USA, at a 1∶300 dilution) at 4°C overnight and then incubated with horseradish peroxidase (HRP) (ChemMateTM DAKO EnVisionTM Detection Kit) at room temperature for 30 min. After washing in PBS, the visualization signal was developed with 3, 3′-diaminobenzidine (DAB) solution, and all of the slides were counterstained with hematoxylin.

Techniques: Western Blot, Expressing, Proliferation Assay, In Vitro