Journal: Scientific Reports

Article Title: A fixation method for the optimisation of western blotting

doi: 10.1038/s41598-019-43039-3

Figure Lengend Snippet: Fixation-dependent differences in lectin staining intensities when using PVDF and nitrocellulose membranes. Pooled human serum proteins (3 μg) were separated on 10% SDS-PAGE, and the proteins were transferred onto PVDF and nitrocellulose membranes, the whole membrane was cutted into five pieces for subsequently fixation treatments and followed by staining with lectins (LCA and SNA). Lane i, CBB staining; lane ii, no fixation; lane iii, drying at room temperature; lane iv, sample heating at 100 °C; lane v, organic solvent (acetone and 50% methanol for PVDF and nitrocellulose membranes, respectively) treatments at room temperature; lane vi, organic solvent treatments followed by sample heating at 100 °C. All treatments were applied for 30 min. Left, WB pattern; right, quantitative analysis (n = 3 individual experiments). The exposure times were the same in all procedures. Band intensities were analysed and compared using Image Lab software (Bio-Rad Laboratories) and GraphPad Prism version 6. ** Significantly different p < 0.01, *** p < 0.001, **** p < 0.0001. All values are means ± S.E. (error bars).

Article Snippet: Biotinylated lectins, LCA, SNA, PHA-E, PHA-L, and AAL (Supplementary Table ), were purchased from Vector Laboratories Inc. (Burlingame, CA, USA).

Techniques: Staining, SDS Page, Software

Journal: American Journal of Physiology - Heart and Circulatory Physiology

Article Title: Enhanced large conductance K + channel activity contributes to the impaired myogenic response in the cerebral vasculature of Fawn Hooded Hypertensive rats

doi: 10.1152/ajpheart.00636.2013

Figure Lengend Snippet: Comparison of BK-α- and BK-β-subunit expression in cerebral vessel homogenates isolated from FHH and FHH.1BN rats. BK-α- and BK-β-subunits were probed on different blots. The same blots were then reprobed for β-actin. A: representative bands corresponding to the molecular weight of the BK-α-subunit (∼100 kDa), BK-β-subunit (∼25 kDa), and β-actin (∼42 kDa) in cerebral vessels obtained from FHH and FHH.1BN rats (n = 8 animals each; 3 sets of experiments). B: a comparison of the expression of α- and β-subunits. Numbers in parentheses indicate the number of samples studied.

Article Snippet: After transfer, the membrane was blocked with TBS-T buffer containing 20 mM Tris pH 7.5, 150 mM NaCl, 0.05% Tween, and a 5% blocking powder (Bio-Rad) at 4°C for 1 h. The blot was probed with primary antibody against BK α- and β-subunit [1:500 and 1:200, respectively; polyclonal rabbit Anti-K Ca 1.1, to amino acids 1184–1200 and Anti-sloβ1 (KCNMB1); Alomone Labs, Jerusalem, Israel] overnight at 4°C.

Techniques: Expressing, Isolation, Molecular Weight

Journal: American Journal of Physiology - Renal Physiology

Article Title: Furosemide reduces BK-αβ4-mediated K + secretion in mice on an alkaline high-K + diet

doi: 10.1152/ajprenal.00223.2018

Figure Lengend Snippet: Furosemide (Furo) effect on urine pH. Urine pH of wild-type (WT; A) and β4-knockout (β4-KO; B) on day −1, 1, 4, 7, and 11 of control (Ctrl) and Furo treatments or on day −1, 1, 4, and 7 of Furo + acetazolamide (Actz) treatment. *P < 0.05 vs. HK + Ctrl water, #P < 0.05 vs. Day –1, analyzed with two-way repeated-measures ANOVA with a post hoc Holm-Sidak test (P < 0.001 for both treatment and days with interaction P < 0.01). n = 12–14 for WT Ctrl and Furo; N = 7 – 8 for β4-KO Ctrl and Furo; N = 4 for WT Furo + Actz; n = 3 for β4-KO Furo + Actz.

Article Snippet: Primary antibodies included anti-BK-β4 (rabbit polyclonal, diluted 1:500; Alomone Laboratories, Jerusalem, Israel), anti-NHE3 (mouse monoclonal, diluted 1:200; Invitrogen, Carlsbad, CA), and anti-V-ATPase B1 (rabbit polyclonal, diluted 1:200; GeneTex, Irvine, CA).

Techniques: Knock-Out

Journal: American Journal of Physiology - Renal Physiology

Article Title: Furosemide reduces BK-αβ4-mediated K + secretion in mice on an alkaline high-K + diet

doi: 10.1152/ajprenal.00223.2018

Figure Lengend Snippet: Blood measurements after 11 days (or 7 days for HK + Furo Actz) of treatment

Article Snippet: Primary antibodies included anti-BK-β4 (rabbit polyclonal, diluted 1:500; Alomone Laboratories, Jerusalem, Israel), anti-NHE3 (mouse monoclonal, diluted 1:200; Invitrogen, Carlsbad, CA), and anti-V-ATPase B1 (rabbit polyclonal, diluted 1:200; GeneTex, Irvine, CA).

Techniques:

Journal: American Journal of Physiology - Renal Physiology

Article Title: Furosemide reduces BK-αβ4-mediated K + secretion in mice on an alkaline high-K + diet

doi: 10.1152/ajprenal.00223.2018

Figure Lengend Snippet: Furosemide (Furo) effect on plasma [K+] and renal K+ clearance in mice on regular diet (RD) and high-K+ diet (HK). A and B: plasma [K+] and renal K+ clearance of WT on RD on day 11 of control (Ctrl) or Furo treatments. *P < 0.05 vs. RD + Ctrl analyzed with Student’s t-test; n = 4 for Ctrl, n = 6 for Furo. C and D: plasma [K+] and renal K+ clearance of WT and β4-knockout (β4-KO) on day 11 of Ctrl and Furo treatment or day 7 of Furo + acetazolamide (Actz) treatment. *P < 0.05 vs. HK + Ctrl; #P < 0.05 vs. HK + Furo, analyzed with two-way ANOVA with a post hoc Tukey test (P < 0.001 for treatment in plasma [K+]; P < 0.01 for treatment in renal K+ clearance); n = 12 – 14 for WT HK + Ctrl and HK + Furo; n = 7 – 8 for β4-KO HK + Ctrl and HK + Furo; n = 4 for WT HK + Furo Actz; n = 3 for β4-KO HK + Furo Actz.

Article Snippet: Primary antibodies included anti-BK-β4 (rabbit polyclonal, diluted 1:500; Alomone Laboratories, Jerusalem, Israel), anti-NHE3 (mouse monoclonal, diluted 1:200; Invitrogen, Carlsbad, CA), and anti-V-ATPase B1 (rabbit polyclonal, diluted 1:200; GeneTex, Irvine, CA).

Techniques: Knock-Out

Journal: American Journal of Physiology - Renal Physiology

Article Title: Furosemide reduces BK-αβ4-mediated K + secretion in mice on an alkaline high-K + diet

doi: 10.1152/ajprenal.00223.2018

Figure Lengend Snippet: Furosemide (Furo) effect on urinary K+ excretion (UKV̇) normalized to kidney weight in mice on high-K diet (HK). UKV̇ of wild-type (WT; A) and β4-knockout (β4-KO; B) on day −1, 1, 4, 7, and 11 of control (Ctrl) and Furo treatment or on day −1, 1, 4, and 7 of Furo + acetazolamide (Actz) treatment. *P < 0.05 vs. HK + Ctrl; #P < 0.05 vs. day −1, analyzed with two-way repeated measures ANOVA (P < 0.05 for drug treatment in WT) with a post hoc Holm-Sidak test; n = 12–14 for WT HK + Ctrl and HK + Furo; n = 7–8 for β4-KO HK + Ctrl and HK + Furo; n = 4 for WT HK + Furo Actz; n = 3 for β4-KO HK + Furo Actz.

Article Snippet: Primary antibodies included anti-BK-β4 (rabbit polyclonal, diluted 1:500; Alomone Laboratories, Jerusalem, Israel), anti-NHE3 (mouse monoclonal, diluted 1:200; Invitrogen, Carlsbad, CA), and anti-V-ATPase B1 (rabbit polyclonal, diluted 1:200; GeneTex, Irvine, CA).

Techniques: Knock-Out

Journal: American Journal of Physiology - Renal Physiology

Article Title: Furosemide reduces BK-αβ4-mediated K + secretion in mice on an alkaline high-K + diet

doi: 10.1152/ajprenal.00223.2018

Figure Lengend Snippet: Metabolic cage measurements after 11 days (or 7 days for HK + Furo Actz) of treatment

Article Snippet: Primary antibodies included anti-BK-β4 (rabbit polyclonal, diluted 1:500; Alomone Laboratories, Jerusalem, Israel), anti-NHE3 (mouse monoclonal, diluted 1:200; Invitrogen, Carlsbad, CA), and anti-V-ATPase B1 (rabbit polyclonal, diluted 1:200; GeneTex, Irvine, CA).

Techniques:

Journal: American Journal of Physiology - Renal Physiology

Article Title: Furosemide reduces BK-αβ4-mediated K + secretion in mice on an alkaline high-K + diet

doi: 10.1152/ajprenal.00223.2018

Figure Lengend Snippet: Correlation between renal K+ clearance and urine pH in wild-type (WT; A) and β4-knockout (β4-KO; B) on high-K+ diet (HK). ●, HK + control (Ctrl) group; ○, HK + furosemide (Furo) group; ▲, HK + Furo acetazolamide (Actz) group; △, HKCl group.

Article Snippet: Primary antibodies included anti-BK-β4 (rabbit polyclonal, diluted 1:500; Alomone Laboratories, Jerusalem, Israel), anti-NHE3 (mouse monoclonal, diluted 1:200; Invitrogen, Carlsbad, CA), and anti-V-ATPase B1 (rabbit polyclonal, diluted 1:200; GeneTex, Irvine, CA).

Techniques: Knock-Out

Journal: American Journal of Physiology - Renal Physiology

Article Title: Furosemide reduces BK-αβ4-mediated K + secretion in mice on an alkaline high-K + diet

doi: 10.1152/ajprenal.00223.2018

Figure Lengend Snippet: Furosemide (Furo) effect on large conductance Ca2+-activated K+ (BK)-β4 expression in the kidney. A: BK-β4 expression in the kidney medulla of wild-type (WT) on high-K diet (HK) treated with control (Ctrl) or Furo water; n = 4/group. B: BK-β4 expression in the kidney cortex of WT on HK treated with Ctrl vs. Furo water. *P < 0.05 vs. HK + Ctrl analyzed with Student’s t-test; n = 7/group. C: BK-β4 expression in the kidney cortex of WT on HK treated with Ctrl vs. Furo acetazolamide (Actz) water; n = 4/group.

Article Snippet: Primary antibodies included anti-BK-β4 (rabbit polyclonal, diluted 1:500; Alomone Laboratories, Jerusalem, Israel), anti-NHE3 (mouse monoclonal, diluted 1:200; Invitrogen, Carlsbad, CA), and anti-V-ATPase B1 (rabbit polyclonal, diluted 1:200; GeneTex, Irvine, CA).

Techniques: Expressing

Journal: American Journal of Physiology - Renal Physiology

Article Title: Furosemide reduces BK-αβ4-mediated K + secretion in mice on an alkaline high-K + diet

doi: 10.1152/ajprenal.00223.2018

Figure Lengend Snippet: Furosemide concentrations in urine and plasma of WT and BK-β4-KO

Article Snippet: Primary antibodies included anti-BK-β4 (rabbit polyclonal, diluted 1:500; Alomone Laboratories, Jerusalem, Israel), anti-NHE3 (mouse monoclonal, diluted 1:200; Invitrogen, Carlsbad, CA), and anti-V-ATPase B1 (rabbit polyclonal, diluted 1:200; GeneTex, Irvine, CA).

Techniques: