primary antibodies against fancd2  (Bethyl)

Journal: Nature Communications

Article Title: ANP32E drives vulnerability to ATR inhibitors by inducing R-loops-dependent transcription replication conflicts in triple negative breast cancer

doi: 10.1038/s41467-025-59804-0

Figure Lengend Snippet: a Dot blot showing R-loop abundance in tIMEC, tIMEC-A, and tIMEC-A-H1 cells ± RNaseH1. dsDNA used as internal normalizer. Quantification from three biological replicates (mean ± s.e.m.), unpaired two-tailed t -test p values shown. b Representative PLA images showing proximity between ANP32E and R-loops. Scale bar = 10 µm. c PLA images of pS2RNApol II-PCNA proximity as TRCs marker. Scale bar = 10 µm. d Quantification of PLA foci/nucleus from ( b , c ): tIMEC n = 106/104, tIMEC-A n = 85/84, tIMEC-A-H1 n = 72/75. Unpaired two-tailed t -test p values are shown. e Top: FANCD2 immunofluorescence in EdU+ cells (EdU channel excluded for clarity). Bottom: quantification of FANCD2 foci/nucleus in EdU+ (green violins) and EdU– (gray violins) cells; unpaired two-tailed t -test p values shown. Scale bar = 10 µm. f Alkaline comet assay ± VE822 (1.25 µM, 24 h). Top: representative images, comet quantification (OpenComet) , and DNA intensity heatmaps (CometScore) . Bottom: violin plots of comet tail length; 150–300 cells/condition, three biological replicates. Unpaired two-tailed t -test was used for p value calculation. g Left: representative binucleated (BN) cells with micronuclei (MNi) after Cytochalasin B + Mitomycin C treatment. Right: %BN cells with MNi ± Mitomycin C (0.03 µg/ml) or VE822 (0.027, 0.08, 0.25 µM) treatment. ≥3000 cells/condition on three biological replicates. The barplot reports mean ± s.d. with two-way ANOVA adjusted p values. h Immunostaining of BN cells with MNi that are positive or negative for centromere staining (CREST) after VE822. Phalloidin marks cytoplasm; arrows indicate MNi. Scale bar = 10 µm. i Barplot showing the percentage of BN cells with DSB-derived (acentric) MNi ± Mitomycin C or VE822 treatment. Mean ± s.e.m. from three replicates; 25–60 BN cells/condition were quantified per replicate.

Article Snippet: Primary antibodies against FANCD2 1:100 (NovusBio, NB100-182SS), pRPA32 1:1000 (Bethyl, A300-246A-8), and 53BP1 1:100 (Millipore, MAB3802) were diluted in blocking solution and incubated at RT for 2 h. Next, coverslips were washed in PBS 1x before incubation with Alexa-Fluor-647 specie-specific secondary antibodies (Thermo Fisher, A32728 and A-21245) and either Hoechst 1:2000 for EdU labeled cells or DAPI 1:1000 for simple IF.

Techniques: Dot Blot, Two Tailed Test, Marker, Immunofluorescence, Alkaline Single Cell Gel Electrophoresis, Immunostaining, Staining, Derivative Assay

Journal: Oncology Letters

Article Title: Expression and prognostic significance of Fanconi anemia group D2 protein and breast cancer type 1 susceptibility protein in familial and sporadic breast cancer

doi: 10.3892/ol.2019.10046

Figure Lengend Snippet: Representative images of BRCA1 and FANCD2 staining in benign and malignant tumor tissues of patients with BC at ×200 magnification. (A) Negative control of BRCA1 in BC tissues. (B) Negative control of FANCD2 in BC tissue. Positive control of BRCA1 in (C) the nuclei and (D) cytoplasm of BC tissue. (E) Positive control of FANCD2 in the nuclei of BC tissues. (F) Positive expression of BRCA1 in the nuclei, (G) BRCA1 in the cytoplasm and (H) FANCD2 in the nuclei of BC tissues. Positive expression of (I) BRCA1 in the nuclei, (J) BRCA1 in the cytoplasm and (K) FANCD2 in the nuclei of the tissues adjacent to the carcinoma. Positive expression of (L) BRCA1 in the nuclei, (M) BRCA1 in the cytoplasm and (N) FANCD2 in the nuclei of benign tissues. (O) Negative expression of FANCD2 in benign tissue. BC, breast cancer; BRCA1, breast cancer type 1 susceptibility protein; FANCD2 Fanconi anemia group D2 protein.

Article Snippet: Membranes were blocked with 5% fat-free milk diluted with TBST (10 mM Tris-HCl, Ph 7.4, 150 mM NaCl, 0.1% Tween-20) at room temperature for 1 h and incubated for 1 h with the rabbit anti-human primary antibodies against FANCD2 (cat. no. sc-28194; 1:1,000) and β-actin (cat. no. sc-47778; 1:3,000) (both Santa Cruz Biotechnology).

Techniques: Staining, Negative Control, Positive Control, Expressing

Journal: Oncology Letters

Article Title: Expression and prognostic significance of Fanconi anemia group D2 protein and breast cancer type 1 susceptibility protein in familial and sporadic breast cancer

doi: 10.3892/ol.2019.10046

Figure Lengend Snippet: Expression of BRCA1 and FANCD2, and clinical characteristics of patients with BC.

Article Snippet: Membranes were blocked with 5% fat-free milk diluted with TBST (10 mM Tris-HCl, Ph 7.4, 150 mM NaCl, 0.1% Tween-20) at room temperature for 1 h and incubated for 1 h with the rabbit anti-human primary antibodies against FANCD2 (cat. no. sc-28194; 1:1,000) and β-actin (cat. no. sc-47778; 1:3,000) (both Santa Cruz Biotechnology).

Techniques: Expressing

Journal: Oncology Letters

Article Title: Expression and prognostic significance of Fanconi anemia group D2 protein and breast cancer type 1 susceptibility protein in familial and sporadic breast cancer

doi: 10.3892/ol.2019.10046

Figure Lengend Snippet: Comparisons of (A) BRCA1 and (B) FANCD2 expression levels between breast cancer tissues and non-cancerous adjacent tissues. The red bar represents the tumor tissues and the gray bar indicates the non-cancerous adjacent tissues. These figures were derived from the Gene Expression Profiling Interactive Analysis dataset. *P<0.05. BRCA1, breast cancer type 1 susceptibility protein; FANCD2 Fanconi anemia group D2 protein.

Article Snippet: Membranes were blocked with 5% fat-free milk diluted with TBST (10 mM Tris-HCl, Ph 7.4, 150 mM NaCl, 0.1% Tween-20) at room temperature for 1 h and incubated for 1 h with the rabbit anti-human primary antibodies against FANCD2 (cat. no. sc-28194; 1:1,000) and β-actin (cat. no. sc-47778; 1:3,000) (both Santa Cruz Biotechnology).

Techniques: Expressing, Derivative Assay, Gene Expression

Journal: Oncology Letters

Article Title: Expression and prognostic significance of Fanconi anemia group D2 protein and breast cancer type 1 susceptibility protein in familial and sporadic breast cancer

doi: 10.3892/ol.2019.10046

Figure Lengend Snippet: Expression of BRCA1 and FANCD2, and clinical characteristics of patients with breast cancer obtained from The Cancer Genome Atlas dataset.

Article Snippet: Membranes were blocked with 5% fat-free milk diluted with TBST (10 mM Tris-HCl, Ph 7.4, 150 mM NaCl, 0.1% Tween-20) at room temperature for 1 h and incubated for 1 h with the rabbit anti-human primary antibodies against FANCD2 (cat. no. sc-28194; 1:1,000) and β-actin (cat. no. sc-47778; 1:3,000) (both Santa Cruz Biotechnology).

Techniques: Expressing

Journal: Oncology Letters

Article Title: Expression and prognostic significance of Fanconi anemia group D2 protein and breast cancer type 1 susceptibility protein in familial and sporadic breast cancer

doi: 10.3892/ol.2019.10046

Figure Lengend Snippet: DFS of patients with breast cancer based on BRCA1 and FANCD2 expression. DFS of patients with FBC grouped by positive or negative expression of (A) BRCA1 and (B) FANCD2. DFS of patients with SBC were divided by positive or negative expression of (C) BRCA1 and (D) FANCD2. Probabilities of DFS were estimated using the Kaplan-Meier method and compared using the log-rank statistic. BRCA1, breast cancer type 1 susceptibility protein; FANCD2, Fanconi anemia group D2 protein; Cum, cumulative; DFS, disease-free survival; FBC, familial breast cancer; SBC, sporadic breast cancer.

Article Snippet: Membranes were blocked with 5% fat-free milk diluted with TBST (10 mM Tris-HCl, Ph 7.4, 150 mM NaCl, 0.1% Tween-20) at room temperature for 1 h and incubated for 1 h with the rabbit anti-human primary antibodies against FANCD2 (cat. no. sc-28194; 1:1,000) and β-actin (cat. no. sc-47778; 1:3,000) (both Santa Cruz Biotechnology).

Techniques: Expressing

Journal: Oncology Letters

Article Title: Expression and prognostic significance of Fanconi anemia group D2 protein and breast cancer type 1 susceptibility protein in familial and sporadic breast cancer

doi: 10.3892/ol.2019.10046

Figure Lengend Snippet: Univariable and multivariable analyses of disease-free survival in the two groups of patients with breast cancer.

Article Snippet: Membranes were blocked with 5% fat-free milk diluted with TBST (10 mM Tris-HCl, Ph 7.4, 150 mM NaCl, 0.1% Tween-20) at room temperature for 1 h and incubated for 1 h with the rabbit anti-human primary antibodies against FANCD2 (cat. no. sc-28194; 1:1,000) and β-actin (cat. no. sc-47778; 1:3,000) (both Santa Cruz Biotechnology).

Techniques:

Journal: Oncology Letters

Article Title: Expression and prognostic significance of Fanconi anemia group D2 protein and breast cancer type 1 susceptibility protein in familial and sporadic breast cancer

doi: 10.3892/ol.2019.10046

Figure Lengend Snippet: Western blot analysis of FANCD2 in patients with SBC. FANCD2 is represented by two adjacent bands. The upper stripe, L, is ubiquitinated FANCD2, and the lower stripe, S, is unubiquitinated FANCD2. Western blotting from the other 50 patients are shown in . BRCA1, breast cancer type 1 susceptibility protein; FANCD2, Fanconi anemia group D2 protein.

Article Snippet: Membranes were blocked with 5% fat-free milk diluted with TBST (10 mM Tris-HCl, Ph 7.4, 150 mM NaCl, 0.1% Tween-20) at room temperature for 1 h and incubated for 1 h with the rabbit anti-human primary antibodies against FANCD2 (cat. no. sc-28194; 1:1,000) and β-actin (cat. no. sc-47778; 1:3,000) (both Santa Cruz Biotechnology).

Techniques: Western Blot

Journal: Oncology Letters

Article Title: Expression and prognostic significance of Fanconi anemia group D2 protein and breast cancer type 1 susceptibility protein in familial and sporadic breast cancer

doi: 10.3892/ol.2019.10046

Figure Lengend Snippet: Distribution of ubiquitination levels of patients with SBC. The L/S ratio of 56 patients with SBC ranged from 0.13 to 1.36, with a median ratio of 0.645. Cases were divided into FANCD2 High and FANCD2 Low groups according to the median L/S ratio of 0.645 (28 cases in each group). FANCD2, Fanconi anemia group D2 protein; SBC, sporadic breast cancer; L/S, ubiquitinated/unubiquitinated.

Article Snippet: Membranes were blocked with 5% fat-free milk diluted with TBST (10 mM Tris-HCl, Ph 7.4, 150 mM NaCl, 0.1% Tween-20) at room temperature for 1 h and incubated for 1 h with the rabbit anti-human primary antibodies against FANCD2 (cat. no. sc-28194; 1:1,000) and β-actin (cat. no. sc-47778; 1:3,000) (both Santa Cruz Biotechnology).

Techniques: Ubiquitin Proteomics

Journal: Oncology Letters

Article Title: Expression and prognostic significance of Fanconi anemia group D2 protein and breast cancer type 1 susceptibility protein in familial and sporadic breast cancer

doi: 10.3892/ol.2019.10046

Figure Lengend Snippet: DFS of patients with SBC based on the ubiquitination level of FANCD2. DFS of patients divided into FANCD2 High and FANCD2 Low groups in the SBC group. Probabilities of DFS were estimated using the Kaplan-Meier method and compared using the log-rank statistic. DFS, disease-free survival; FANCD2, Fanconi anemia group D2 protein; SBC, sporadic breast cancer.

Article Snippet: Membranes were blocked with 5% fat-free milk diluted with TBST (10 mM Tris-HCl, Ph 7.4, 150 mM NaCl, 0.1% Tween-20) at room temperature for 1 h and incubated for 1 h with the rabbit anti-human primary antibodies against FANCD2 (cat. no. sc-28194; 1:1,000) and β-actin (cat. no. sc-47778; 1:3,000) (both Santa Cruz Biotechnology).

Techniques: Ubiquitin Proteomics

Journal: Oncology Letters

Article Title: Expression and prognostic significance of Fanconi anemia group D2 protein and breast cancer type 1 susceptibility protein in familial and sporadic breast cancer

doi: 10.3892/ol.2019.10046

Figure Lengend Snippet: Univariable and multivariable analyses of disease-free survival in patients with sporadic breast cancer (n=141).

Article Snippet: Membranes were blocked with 5% fat-free milk diluted with TBST (10 mM Tris-HCl, Ph 7.4, 150 mM NaCl, 0.1% Tween-20) at room temperature for 1 h and incubated for 1 h with the rabbit anti-human primary antibodies against FANCD2 (cat. no. sc-28194; 1:1,000) and β-actin (cat. no. sc-47778; 1:3,000) (both Santa Cruz Biotechnology).

Techniques: Ubiquitin Proteomics