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pcna  (Cell Signaling Technology Inc)
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Cell Signaling Technology Inc pcna
(A) Protein blots of samples harvested 72 h after silencing ferritin light <t>chain</t> <t>(FTL)</t> in A172 glioblastoma multiforme (GBM) cells with three respective si-RNAs: si-FTL-1, si-FTL-2, and si-FTL-3. Of these, si-FTL-2 showed the best interference efficiency (si-control group vs. si-FTL-2 group: P < 0.01). “Mock” denotes treatment with transfection reagent only. “si-Control” denotes transfection with scrambled si-RNA as a negative control. GAPDH was used as an internal loading control. The number at the top of each band represents the average densitometric value normalized against that of GAPDH. (B–D) Results of Cell Counting Kit-8 (CCK-8) assays 24, 48, and 72 h after transfection of GBM cells with mock, si-FTL-2, or scrambled control (negative control: NC) si-RNAs. The data points represent the OD ratio of Mock/NC, NC/NC, si-FTL-2/NC at different time points. Data represent the mean ± standard deviation of six replicates per time point. (E) Immunoblot analyses of U251, A172, and U87 GBM cells. Downregulation of FTL resulted in reduced expression of topoisomerase II-alpha (Topo II) and p-Histone H3 (Ser10) (p-His3) in all cell lines (P < 0.01). <t>PCNA</t> expression also decreased in A172 and U87 cells (P < 0.01), but not in U251 cells (p = 0.04). GAPDH was used as an internal loading control. The number at the top of each band represents the average densitometric value from three experiments, normalized against that of GAPDH (FTL, Topo II and PCNA) or His3 (p-His3).
Pcna, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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(A) Protein blots of samples harvested 72 h after silencing ferritin light chain (FTL) in A172 glioblastoma multiforme (GBM) cells with three respective si-RNAs: si-FTL-1, si-FTL-2, and si-FTL-3. Of these, si-FTL-2 showed the best interference efficiency (si-control group vs. si-FTL-2 group: P < 0.01). “Mock” denotes treatment with transfection reagent only. “si-Control” denotes transfection with scrambled si-RNA as a negative control. GAPDH was used as an internal loading control. The number at the top of each band represents the average densitometric value normalized against that of GAPDH. (B–D) Results of Cell Counting Kit-8 (CCK-8) assays 24, 48, and 72 h after transfection of GBM cells with mock, si-FTL-2, or scrambled control (negative control: NC) si-RNAs. The data points represent the OD ratio of Mock/NC, NC/NC, si-FTL-2/NC at different time points. Data represent the mean ± standard deviation of six replicates per time point. (E) Immunoblot analyses of U251, A172, and U87 GBM cells. Downregulation of FTL resulted in reduced expression of topoisomerase II-alpha (Topo II) and p-Histone H3 (Ser10) (p-His3) in all cell lines (P < 0.01). PCNA expression also decreased in A172 and U87 cells (P < 0.01), but not in U251 cells (p = 0.04). GAPDH was used as an internal loading control. The number at the top of each band represents the average densitometric value from three experiments, normalized against that of GAPDH (FTL, Topo II and PCNA) or His3 (p-His3).

Journal: PLoS ONE

Article Title: Expression of Ferritin Light Chain (FTL) Is Elevated in Glioblastoma, and FTL Silencing Inhibits Glioblastoma Cell Proliferation via the GADD45/JNK Pathway

doi: 10.1371/journal.pone.0149361

Figure Lengend Snippet: (A) Protein blots of samples harvested 72 h after silencing ferritin light chain (FTL) in A172 glioblastoma multiforme (GBM) cells with three respective si-RNAs: si-FTL-1, si-FTL-2, and si-FTL-3. Of these, si-FTL-2 showed the best interference efficiency (si-control group vs. si-FTL-2 group: P < 0.01). “Mock” denotes treatment with transfection reagent only. “si-Control” denotes transfection with scrambled si-RNA as a negative control. GAPDH was used as an internal loading control. The number at the top of each band represents the average densitometric value normalized against that of GAPDH. (B–D) Results of Cell Counting Kit-8 (CCK-8) assays 24, 48, and 72 h after transfection of GBM cells with mock, si-FTL-2, or scrambled control (negative control: NC) si-RNAs. The data points represent the OD ratio of Mock/NC, NC/NC, si-FTL-2/NC at different time points. Data represent the mean ± standard deviation of six replicates per time point. (E) Immunoblot analyses of U251, A172, and U87 GBM cells. Downregulation of FTL resulted in reduced expression of topoisomerase II-alpha (Topo II) and p-Histone H3 (Ser10) (p-His3) in all cell lines (P < 0.01). PCNA expression also decreased in A172 and U87 cells (P < 0.01), but not in U251 cells (p = 0.04). GAPDH was used as an internal loading control. The number at the top of each band represents the average densitometric value from three experiments, normalized against that of GAPDH (FTL, Topo II and PCNA) or His3 (p-His3).

Article Snippet: The primary antibodies used in this study recognized the following proteins: human FTL (Abcam, Cambridge, MA, USA: Ab109019; 1:1000), PCNA (Cell Signaling Technology, Danvers, MA, USA: 13110; 1:1000), topoisomerase II-alpha (Abcam: Ab2987; 1:100), GAPDH (Santa Cruz Biotechnology, CA, USA: sc-25778; 1:1000), p-Histone H3 (Ser10) (Santa Cruz Biotechnology: sc-8656-R; 1:1000), Histone H3 (Cell Signaling Technology: 4499; 1:1000), GADD45a (Santa Cruz Biotechnology: sc-6850; 1:200), Cyclin D1 (Cell Signaling Technology: 2978; 1:1000), c-Myc (Santa Cruz Biotechnology: sc-40; 1:1000), PDGFR-β (Santa Cruz Biotechnology: sc-432; 1:1000), p-JNK (Santa Cruz Biotechnology: sc-6254; 1:1000), JNK (Santa Cruz Biotechnology: sc-7345; 1:1000), p-Histone H2A.X (Ser139) (Cell Signaling Technology: 9718; 1:1000), and Histone H2A (Cell Signaling Technology: 12349; 1:1000).

Techniques: Control, Transfection, Negative Control, Cell Counting, CCK-8 Assay, Standard Deviation, Western Blot, Expressing