primary antibodies for terf2  (Novus Biologicals)

Journal: Translational Cancer Research

Article Title: High TERF2 expression is associated with poor prognosis and its suppression attenuates progression in acute myeloid leukemia

doi: 10.21037/tcr-2025-1226

Figure Lengend Snippet: Elevated TERF2 expression in AML patients and its correlation with clinicopathological features. (A) Transcriptional expression of TERF2 in AML analyzed using TCGA database. (B) TERF2 mRNA levels quantified by qRT-PCR in PBMCs from 50 AML patients and 35 healthy donors. (C) UMAP of single cells from an AML patient ( GSE116256 ), single-cell profiling reveals distinct expression patterns of TERF2 across cellular subpopulations in AML. (D) Heatmap depicting the average expression level of TERF2 across distinct cell types within the AML sample. (E) Univariate Cox proportional hazards regression analysis comparing survival outcomes between high- and low-TERF2 expression groups. (F) Prognostic accuracy of TERF2 evaluated using ROC curve analysis. (G) The predictive performance of the novel risk stratification model was evaluated through time-dependent ROC curve analysis, with AUC values calculated at 1-, 3-, and 5-year intervals to quantify sensitivity and specificity in AML prognosis. ***, P<0.001; ****, P<0.0001. AML, acute myeloid leukemia; AUC, area under the curve; CI, confidence interval; FPR, false positive rate; HR, hazard ratio; PBMCs, peripheral blood mononuclear cells; qRT-PCR, quantitative reverse transcription polymerase chain reaction; ROC, receiver operating characteristic; TCGA, The Cancer Genome Atlas; TPM, transcripts per million; TPR, true positive rate; UMAP, uniform manifold approximation and projection.

Article Snippet: Primary antibodies targeting TERF2 (#13136, 1:500 dilution), CDK4 (#12790, 1:500), CDK6 (#1331, 1:500), and CDKN2A (#80772, 1:500) were commercially obtained from CST (Danvers, MA, USA).

Techniques: Expressing, Quantitative RT-PCR, Reverse Transcription, Polymerase Chain Reaction

Journal: Translational Cancer Research

Article Title: High TERF2 expression is associated with poor prognosis and its suppression attenuates progression in acute myeloid leukemia

doi: 10.21037/tcr-2025-1226

Figure Lengend Snippet: Downregulation of TERF2 suppresses AML cells viability and proliferation. (A) qRT-PCR assays analyzed TERF2 mRNA levels of NC or TERF2-knockdown MOLM13 cells and THP1 cells. (B) Western blot assays detected TERF2 protein levels of NC or TERF2-knockdown MOLM13 cells and THP1 cells. (C,D) CCK-8 assay showing the viability of NC or TERF2-knockdown MOLM13 cells (C) and THP1 cells (D). (E-H) Flow cytometry assay analyzed cell cycle in NC or TERF2-knockdown MOLM13 cells (E,F) and THP1 cells (G,H). **, P<0.01; ****, P<0.0001; ns, not significant. AML, acute myeloid leukemia; CCK-8, Cell Counting Kit-8; NC, negative control; qRT-PCR, quantitative reverse transcription polymerase chain reaction.

Article Snippet: Primary antibodies targeting TERF2 (#13136, 1:500 dilution), CDK4 (#12790, 1:500), CDK6 (#1331, 1:500), and CDKN2A (#80772, 1:500) were commercially obtained from CST (Danvers, MA, USA).

Techniques: Quantitative RT-PCR, Knockdown, Western Blot, CCK-8 Assay, Flow Cytometry, Cell Counting, Negative Control, Reverse Transcription, Polymerase Chain Reaction

Journal: Translational Cancer Research

Article Title: High TERF2 expression is associated with poor prognosis and its suppression attenuates progression in acute myeloid leukemia

doi: 10.21037/tcr-2025-1226

Figure Lengend Snippet: TERF2 deficiency promotes apoptosis in AML cells. (A-D) Flow cytometry assay analyzed apoptosis in NC or TERF2-knockdown MOLM13 cells (A,B) or THP1 cells (C,D). (E-H) Western blot detected cleaved caspase 3 levels of NC or TERF2-knockdown MOLM13 cells (E,F) or THP1 cells (G,H). ***, P<0.001; ****, P<0.0001. AML, acute myeloid leukemia; NC, negative control.

Article Snippet: Primary antibodies targeting TERF2 (#13136, 1:500 dilution), CDK4 (#12790, 1:500), CDK6 (#1331, 1:500), and CDKN2A (#80772, 1:500) were commercially obtained from CST (Danvers, MA, USA).

Techniques: Flow Cytometry, Knockdown, Western Blot, Negative Control

Journal: Translational Cancer Research

Article Title: High TERF2 expression is associated with poor prognosis and its suppression attenuates progression in acute myeloid leukemia

doi: 10.21037/tcr-2025-1226

Figure Lengend Snippet: TERF2 involved in cuproptosis through E2F pathway in AML. (A) Bar graphs illustrate the associations between TERF2 expression levels and gene hallmark sets in AML patients. (B) GSEA for AML patients with high TERF2 expression in TCGA database. (C-E) Western blot analysis of E2F1, CDK4, CDK6, CDKN2A levels in MOLM13 cells and NB4 cells with or without TERF2 silence. (F,G) MOLM13 (F) and NB4 (G) cells with TERF2 knockdown were treated with specified concentrations of ES-Cu (1:1 ratio) for 48 hours, followed by cell viability assessment using the CCK-8 assay. ***, P<0.001; ****, P<0.0001. AML, acute myeloid leukemia; CCK-8, Cell Counting Kit-8; ES-Cu, elesclomol-copper; GSEA, gene set enrichment analysis; IC 50 , half-maximal inhibitory concentration; TCGA, The Cancer Genome Atlas.

Article Snippet: Primary antibodies targeting TERF2 (#13136, 1:500 dilution), CDK4 (#12790, 1:500), CDK6 (#1331, 1:500), and CDKN2A (#80772, 1:500) were commercially obtained from CST (Danvers, MA, USA).

Techniques: Expressing, Western Blot, Knockdown, CCK-8 Assay, Cell Counting, Concentration Assay

Journal: Translational Cancer Research

Article Title: High TERF2 expression is associated with poor prognosis and its suppression attenuates progression in acute myeloid leukemia

doi: 10.21037/tcr-2025-1226

Figure Lengend Snippet: Knockdown of TERF2 suppresses AML progression and enhances cuproptosis sensitivity in vivo . (A,B) Tumor growth was monitored by bioluminescence imaging in MOLM13-engrafted NSG mice (A) and the quantification of luciferase signals for all mice per group (B). (C) Kaplan-Meier analysis of survival of MOLM13-engrafted mice, log rank test. (D,E) Tumor growth was monitored by bioluminescence imaging in MOLM13-engrafted mice treated with elesclomol (D) and the quantification of luciferase signals for all mice per group (E). (F) Kaplan-Meier analysis of survival of MOLM13-engrafted mice treated with elesclomol, log rank test. **, P<0.01. AML, acute myeloid leukemia; NSG, NOD-SCID IL2rg.

Article Snippet: Primary antibodies targeting TERF2 (#13136, 1:500 dilution), CDK4 (#12790, 1:500), CDK6 (#1331, 1:500), and CDKN2A (#80772, 1:500) were commercially obtained from CST (Danvers, MA, USA).

Techniques: Knockdown, In Vivo, Imaging, Luciferase

Journal: Cell reports

Article Title: Topoisomerase IIIα controls alternative lengthening of telomeres

doi: 10.1016/j.celrep.2025.116066

Figure Lengend Snippet: (A) Representative immunofluorescence images showing colocalization of HALO-TOP3A with GFP-TERF2 in U2OS. The scale bar represents 10 μm. (B) Representative immunofluorescence images showing localization of TOP3A with TERF2 in U2OS (ALT). (C) Quantification for TOP3A-TERF2 co-foci per nucleus in U2OS. 100 nuclei were calculated. (D) Representative immunofluorescence images showing localization of TOP3A with TERF2 in U2OS (ALT) and SJSA1 (Tel) cells. The scale bar represents 20 μm. (E) Quantification for TOP3A-TERF2 co-foci per nucleus in U2OS and SJSA1 cells. 25 nuclei per experiment were calculated and compared ( n = 2). (F) Proximity ligation assay (PLA) showing interaction of TOP3A with TERF2 and PML in U2OS cells. The scale bar represents 10 μm.

Article Snippet: Primary antibodies for TERF2 (1:1000 dilution, NB110–57130, Novus Biologicals, rabbit and 1:250 dilution,4A794, Millipore sigma, mouse), PML (1:250 dilution, sc-966, Santa Cruz Biotechnology, mouse) and BLM (1:250 dilution, A-300–110, Bethyl laboratories, Rabbit) in the blocking buffer were incubated for 1 h at room temperature.

Techniques: Immunofluorescence, Proximity Ligation Assay

Journal: Cell reports

Article Title: Topoisomerase IIIα controls alternative lengthening of telomeres

doi: 10.1016/j.celrep.2025.116066

Figure Lengend Snippet: (A) Representative immunofluorescence images showing localization of PML and TERF2 in U2OS cells after siTOP3A treatment. The scale bar represents 10 μm. (B) Quantification for TERF2-PML co-foci per nucleus. 40 nuclei were counted for each condition ( n = 3). Data are plotted as means ± SDs. **** p ≤ 0.0001 (two-tailed unpaired t test). (C) Representative immunofluorescence images showing localization of BLM and TERF2 in U2OS cells after siTOP3A treatment. The scale bar represents 10 μm. (D) Quantification for TERF2-BLM co-foci per nucleus. 40 nuclei were counted for each condition ( n = 3). Error bars represent means ± SDs. **** p ≤ 0.0001 (two-tailed unpaired t test). (E) Schematic representation of ssTeloC detection (telomere probe under native conditions). (F) Representative images of ssTeloC staining performed after native FISH (scale bar: 10 μm). (G) Quantification for ssTeloC foci per nucleus in U2OS. 50 nuclei were counted for each condition ( n = 3). Data are plotted as means ± SDs. **** p ≤ 0.0001 (two-tailed unpaired t test).

Article Snippet: Primary antibodies for TERF2 (1:1000 dilution, NB110–57130, Novus Biologicals, rabbit and 1:250 dilution,4A794, Millipore sigma, mouse), PML (1:250 dilution, sc-966, Santa Cruz Biotechnology, mouse) and BLM (1:250 dilution, A-300–110, Bethyl laboratories, Rabbit) in the blocking buffer were incubated for 1 h at room temperature.

Techniques: Immunofluorescence, Two Tailed Test, Staining

Journal: Cell reports

Article Title: Topoisomerase IIIα controls alternative lengthening of telomeres

doi: 10.1016/j.celrep.2025.116066

Figure Lengend Snippet: (A) Scheme of TERRA R-loops. (B) Representative images of RNA FISH for TERRA in U2OS cells after TOP3A knockdown (scale bar: 10 μm). (C) Quantification of TERRA foci per nucleus. 50 nuclei were counted for each condition ( n = 3). Error bars represent means ± SDs. **** p ≤ 0.0001 (two-tailed unpaired t test). (D) Quantification for TERF2 foci per nucleus in U2OS. 40 nuclei were counted for each condition ( n = 3). Data are plotted as means ± SDs. **** p ≤ 0.0001 (two-tailed unpaired t test).

Article Snippet: Primary antibodies for TERF2 (1:1000 dilution, NB110–57130, Novus Biologicals, rabbit and 1:250 dilution,4A794, Millipore sigma, mouse), PML (1:250 dilution, sc-966, Santa Cruz Biotechnology, mouse) and BLM (1:250 dilution, A-300–110, Bethyl laboratories, Rabbit) in the blocking buffer were incubated for 1 h at room temperature.

Techniques: Knockdown, Two Tailed Test

Journal: Cell reports

Article Title: Topoisomerase IIIα controls alternative lengthening of telomeres

doi: 10.1016/j.celrep.2025.116066

Figure Lengend Snippet: (A) Fluorescence-activated cell sorting (FACS) analysis of HT1080 cells transfected with siTOP3A and harvested after 72 h. EdU (10 μM) was added during the final 30 min before cell harvesting. EdU/DAPI uptake per cell was analyzed. (B) FACS analysis of U2OS cells transfected with siTOP3A and harvested after 72 h. EdU (10 μM) was added during the final 30 min before cell harvesting. EdU/DAPI uptake per cell was analyzed. (C) Representative western blots after TOP3A knockdown in U2OS and HT1080 cells. (D) Quantification of TERF2 intensity using ImageJ software and normalization with GAPDH controls. The experiment was performed in triplicate and the average TERF2 intensity calculated. Error bars represent means ± SDs. n.s., not significant and *** p ≤ 0.0008 (two-tailed unpaired t test). (E) Cycloheximide chase experiment monitoring TERF2 stability after siRNA-mediated TOP3A knockdown for 48 h in U2OS cells at the indicated time intervals. (F) Quantification of TERF2 intensity in U2OS using ImageJ software and normalization with GAPDH. (G) Cycloheximide chase experiment monitoring TERF2 stability after siRNA-mediated TOP3A knockdown for 72 h in HT1080 cells at the indicated time intervals. (H) Quantification of TERF2 intensity in HT1080 using ImageJ software and normalization with PCNA.

Article Snippet: Primary antibodies for TERF2 (1:1000 dilution, NB110–57130, Novus Biologicals, rabbit and 1:250 dilution,4A794, Millipore sigma, mouse), PML (1:250 dilution, sc-966, Santa Cruz Biotechnology, mouse) and BLM (1:250 dilution, A-300–110, Bethyl laboratories, Rabbit) in the blocking buffer were incubated for 1 h at room temperature.

Techniques: Fluorescence, FACS, Transfection, Cell Harvesting, Western Blot, Knockdown, Software, Two Tailed Test

Journal: Cell reports

Article Title: Topoisomerase IIIα controls alternative lengthening of telomeres

doi: 10.1016/j.celrep.2025.116066

Figure Lengend Snippet: (A) Domain organization of human TOP3A. The arrow indicates the catalytic tyrosine and the adjacent arginine residue. (B) Representative images of RNA FISH for TERRA in U2OS cells after overexpression of WT TOP3A and TOP3A R364W. Scale bar: 20 μm. (C) Quantification for TERRA foci per nucleus in U2OS cells after overexpression of WT TOP3A and TOP3A R364W. In total, 50 nuclei were calculated ( n = 2). Data are plotted as means ± SDs. **** p ≤ 0.0001 (two-tailed unpaired t test). (D) Representative TERF2 immunoblot after overexpression of WT TOP3A and TOP3A R364W.

Article Snippet: Primary antibodies for TERF2 (1:1000 dilution, NB110–57130, Novus Biologicals, rabbit and 1:250 dilution,4A794, Millipore sigma, mouse), PML (1:250 dilution, sc-966, Santa Cruz Biotechnology, mouse) and BLM (1:250 dilution, A-300–110, Bethyl laboratories, Rabbit) in the blocking buffer were incubated for 1 h at room temperature.

Techniques: Residue, Over Expression, Two Tailed Test, Western Blot

Journal: Cell reports

Article Title: Topoisomerase IIIα controls alternative lengthening of telomeres

doi: 10.1016/j.celrep.2025.116066

Figure Lengend Snippet: (A) TOP3A interacts with the components of the BTRR complex and TERF2 and protects the shelterin complex from destabilization in ALT cells. (B) TOP3A promotes the formation of TERRA R-loops and c-circles at the telomeres of ALT cells. (C) TOP3A resolves topological DNA structures associated with D-loops during ALT synthesis (steps 1–2); TOP3A can also resolve double Holliday junctions [DHJs] to release extended telomeres without crossover [steps 5–6]) (see for details).

Article Snippet: Primary antibodies for TERF2 (1:1000 dilution, NB110–57130, Novus Biologicals, rabbit and 1:250 dilution,4A794, Millipore sigma, mouse), PML (1:250 dilution, sc-966, Santa Cruz Biotechnology, mouse) and BLM (1:250 dilution, A-300–110, Bethyl laboratories, Rabbit) in the blocking buffer were incubated for 1 h at room temperature.

Techniques: