primary mouse anti β actin mab  (Santa Cruz Biotechnology)

Journal: PLOS One

Article Title: FOXO1 transcription factor modulates airway epithelial responses to viral infection

doi: 10.1371/journal.pone.0345169

Figure Lengend Snippet: A) RT-qPCR showed increased TLR3 mRNA expression for BEAS-2B transfected with a CA-FOXO1 plasmid compared to vector control (cells transfected with an empty plasmid); GAPDH was used as a housekeeping gene (n = 6). Representative Western blot (B) and densitometry analysis (C) of TLR3 expression for BEAS-2B transfected with CA-FOXO1 plasmid compared to vector control, β-actin was used as a loading control (n = 6). Statistical Analysis with t-test, **p < 0.01. D + E) Immunofluorescence staining for BEAS-2B transduced with CA-FOXO1 shows increased FOXO1 protein in the nucleus. FOXO1 (red) was detected using an anti-FOXO1 antibody with a red-fluorescent secondary antibody, F-actin (green) with phalloidin, and nuclei (blue) with DAPI. Images were taken with an Olympus IX81 epifluorescence microscope using a 20X objective lens. Volocity Analysis was used to quantify nuclear localization of FOXO1 by measuring the mean fluorescence intensity of FOXO1 staining colocalized with DAPI. For each group 40−60 cells per slide were analyzed. Statistical Analysis was conducted with ANOVA **** p < 0.001. BEAS-2B cells transduced with FOXO1 or scrambled shRNA lentivirus were analyzed by RT-qPCR for DDX58 (RIG-I, F), MAVS (G), and MYD88 (H) mRNA expression at baseline and after Poly(I:C) stimulation (8 h and 24 h). Expression was normalized to GAPDH and expressed relative to unstimulated scrambled controls (n = 3; ANOVA). (I) NHBE cells were infected with SARS-CoV-2 in the presence or absence of a FOXO1 inhibitor. Total RNA was collected 24 h post-infection, and viral RNA levels were quantified by qRT-PCR, normalized to ACTB, and expressed relative to mock-infected cells (n = 3; paired t-test).

Article Snippet: Primary mouse anti-β-actin mAb (Santa Cruz Biotechnology, SC-69679) and IRdye-conjugated donkey anti-mouse IgG (LI-COR, Lincoln, Neb) were used as a loading control.

Techniques: Quantitative RT-PCR, Expressing, Transfection, Plasmid Preparation, Control, Western Blot, Immunofluorescence, Staining, Transduction, Microscopy, Fluorescence, shRNA, Infection

Journal: Chemmedchem

Article Title: Impact of C18 Epimerization of Indole‐ and Pyrazole‐Fused 18β‐Glycyrrhetinic Acid Derivatives on PTP1B and TCPTP Inhibitory Activity: Synthesis, In Vitro, and In Silico Studies

doi: 10.1002/cmdc.202500350

Figure Lengend Snippet: Effects of metformin, ursolic acid, FC‐114 , FC ‐ 122 , 3c , and 5c on the levels of PTP1B and pAKT in an insulin resistance model in HepG2 cells. a) Western blotting representative analysis ( n = 3) of PTP1B and pAKT levels of HepG2 cells treated with the compounds at 24 h of treatment. β‐actin was employed as the loading control. b) Densitometric analysis of western blot images employing ImageJ analysis software. Bar graphs represent the mean ± SEM of normalized values. The control levels were set at 1. T‐test significance * p < 0.05, when compared with control (DMSO). Statistical analyses were carried out using GraphPad Prism 8.0.

Article Snippet: Rabbit anti‐PTP1B (5311S) and mouse anti‐ β actin (3700S) primary antibodies were obtained from Cell Signaling, and secondary antibodies coupled with horseradish peroxidase were from Jackson laboratories (rabbit‐111−035, mouse‐115–035).

Techniques: Western Blot, Control, Software

Journal: Fluids and Barriers of the CNS

Article Title: Neuroprotective role of pyrroloquinoline quinone in folate deficiency-induced blood-brain barrier disruption

doi: 10.1186/s12987-025-00689-y

Figure Lengend Snippet: Effects of FD and PQQ treatment on the mRNA expression of folate transporters SLC19A1 (RFC) and SLC46A1 (PCFT) in hCMEC/D3 cells. Cells cultured in a FD or FS control medium were treated with PQQ (1 μM or 5 µM) or vehicle (DMSO) for 24 h. ( A ) SLC19A1 (RFC) and ( B ) SLC46A1 (PCFT) mRNA levels were measured using qPCR. Cyclophilin B was used as the housekeeping gene. Results are presented as mean relative mRNA expression normalized to the DMSO vehicle control ± SEM from n = 4–5 independent experiments using cells from different passages. ( C ) RFC and ( D ) PCFT protein bands were detected by western blot analysis using anti-SLC19A1 and anti-SLC46A1 antibodies. For PCFT, multiple bands were observed, corresponding to different glycosylated forms. β-actin was used as the loading control. Densitometric analysis was performed using Image Lab software (Bio-Rad) to quantify protein band intensities. Protein expression data are presented as mean relative protein expression normalized to the corresponding DMSO vehicle control ± SEM from n = 3 independent experiments using cells from different passages. Statistical analysis was performed using two-way ANOVA with Bonferroni’s post-hoc test. Asterisks represent significant differences. *= p < 0.05, **= p < 0.01, ***= p < 0.001, ****= p < 0.0001

Article Snippet: Primary Mouse monoclonal anti-β-actin antibody (sc-517582) was obtained from Santa Cruz Biotechnology.

Techniques: Expressing, Cell Culture, Control, Western Blot, Software

Journal: Fluids and Barriers of the CNS

Article Title: Neuroprotective role of pyrroloquinoline quinone in folate deficiency-induced blood-brain barrier disruption

doi: 10.1186/s12987-025-00689-y

Figure Lengend Snippet: Effects of FD and PQQ treatment on the gene and protein expression of TJ proteins in hCMEC/D3 cells. Cells cultured in a FD or FS control medium were treated with PQQ (1 μM or 5 µM) or vehicle (DMSO) for 24 h. ( A ) Zonula occludens-1 ( ZO-1/TJP1 ), ( B ) Occludin ( OCLN ) and, ( C ) Claudin-5 ( CLDN5 ) mRNA levels were measured using qPCR. Cyclophilin B was used as the housekeeping gene. Results are presented as mean relative mRNA expression normalized to the DMSO vehicle control ± SEM from n = 4–5 independent experiments using cells from different passages. Protein expression of ( D ) TJP1, ( E ) OCLN, and ( F ) CLDN5 was assessed by western blot analysis using anti-ZO-1, anti-OCLN, and anti-CLDN5 antibodies. β-actin was used as the loading control. Densitometric analysis was performed using Image Lab software (Bio-Rad) to quantify protein band intensities. Protein expression data are presented as mean relative protein expression normalized to the DMSO vehicle control ± SEM from n = 3-4 independent experiments using cells from different passages. Statistical analysis was performed using two-way ANOVA with Bonferroni’s post-hoc test. *= p < 0.05, **= p < 0.01, ***= p < 0.001, ****= p < 0.0001

Article Snippet: Primary Mouse monoclonal anti-β-actin antibody (sc-517582) was obtained from Santa Cruz Biotechnology.

Techniques: Expressing, Cell Culture, Control, Western Blot, Software