primary antibodies anti kcnn3  (Alomone Labs)

Journal: bioRxiv

Article Title: High-resolution integrative analysis allows characterization and spatial annotation of normal and cancer-associated colon fibroblasts

doi: 10.1101/2025.07.29.667377

Figure Lengend Snippet: (A) Dot plot depicting marker genes for mouse MIFs (muscle-embedded interstitial fibroblasts). (B) GSEA of the mouse MIF signature against indicated Gene Ontology Molecular Function (GOMF) gene sets. (C) Immunofluorescent staining of PDGFRA and KCNN3 in the muscularis propria in mouse colon and (D) small intestine. Dashed line indicates crypts. (E) Dot plot depicting marker genes for indicated human cell types. (F) GSEA of human MIF signature against indicated GOMF gene sets. (G) Normalized enrichment scores (NES) of GSEA of human MIF signature against gene sets from human colon-derived cells: PDGFRA+ cells of the smooth muscle (PaC), intestinal cells of Cajal (ICC), PDGFRA+ cells of the colon mucosa (FIB), smooth muscle cells of the colon muscle (SMC), enteric neurons from colon myenteric ganglia (EN). (H) Immunofluorescent staining of PDGFRA and KCNN3 in the muscularis propria in human colon. (I) Graphical representation of MIFs in the mouse and human colon.

Article Snippet: Primary antibodies anti-KCNN3 (APC-025, Alomone Labs, 1:100 dilution) and anti-PDGFRa (AF-307-NA, R&D Systems, 1:50 dilution) were incubated overnight at +4°C.

Techniques: Marker, Staining, Derivative Assay

Journal: PLoS ONE

Article Title: Upregulation of SK3 and IK1 Channels Contributes to the Enhanced Endothelial Calcium Signaling and the Preserved Coronary Relaxation in Obese Zucker Rats

doi: 10.1371/journal.pone.0109432

Figure Lengend Snippet: (A, B) Isometric tension recordings showing the effects of increasing concentrations of acetylcholine (ACh) on coronary arteries precontracted with 5-HT (1–3 µM) and effect of treatment with the SK3 channel inhibitor apamin (0.5 µM) plus the IK1 channel inhibitor TRAM34 (0.4 µM) in LZR (A) and OZR (B). (C–D) Effects of apamin, TRAM34, apamin plus TRAM34 and L-NOARG (100 µM) on the average concentration-dependent curves for the relaxation to ACh in coronary arteries from LZR (C) and OZR (D). Results are expressed as a percentage of the contraction induced by 5-HT. Points represent mean ± SEM of n = 6–9 arteries (1–2 arteries per animal).

Article Snippet: Protein expression was quantified using primary antibodies anti-SK3 (Alomone, Israel, 1∶200 dilution), anti-IK1 (Alomone, Israel, 1∶450 dilution), or anti-β-actin as a loading control (Sigma-Aldrich, Spain, 1∶10000 dilution) and horseradish peroxidase conjugated secondary goat anti-mouse and anti-rabbit antibodies (Santa Cruz Biotech, CA, USA, 1∶10000 dilution).

Techniques: Concentration Assay

Journal: PLoS ONE

Article Title: Upregulation of SK3 and IK1 Channels Contributes to the Enhanced Endothelial Calcium Signaling and the Preserved Coronary Relaxation in Obese Zucker Rats

doi: 10.1371/journal.pone.0109432

Figure Lengend Snippet: Effect of selective inhibitors of NOS (L-NOARG), SK3 (apamin) and IK1 channels (TRAM 34), and combined inhibition of SK3/IK1 channels (A+T) on the ACh and SNAP-induced vasodilation in coronary arteries from LZR and OZR.

Article Snippet: Protein expression was quantified using primary antibodies anti-SK3 (Alomone, Israel, 1∶200 dilution), anti-IK1 (Alomone, Israel, 1∶450 dilution), or anti-β-actin as a loading control (Sigma-Aldrich, Spain, 1∶10000 dilution) and horseradish peroxidase conjugated secondary goat anti-mouse and anti-rabbit antibodies (Santa Cruz Biotech, CA, USA, 1∶10000 dilution).

Techniques: Inhibition

Journal: PLoS ONE

Article Title: Upregulation of SK3 and IK1 Channels Contributes to the Enhanced Endothelial Calcium Signaling and the Preserved Coronary Relaxation in Obese Zucker Rats

doi: 10.1371/journal.pone.0109432

Figure Lengend Snippet: (A) The selective opener of SK3 and IK1 channels NS309 induces a larger relaxant effect on coronary arteries of OZR. (B, C) Effect of endothelium removal and further blockade of SK3/IK1 channels with apamin (0.5 µM) plus TRAM (0.4 µM) on the relaxations induced by NS309 in coronary arteries of LZR (B) and OZR (C). (D, E) Effect of NOS blockade with L-NOARG (100 µM) and further inhibition of SK3/IK1 channels with apamin plus TRAM on the relaxations elicited by NS309 in coronary arteries of LZR (D) and OZR (E). Results are expressed as a percentage of the contraction induced by serotonin. Points represent mean ± SEM of n = 9–13 arteries (1–2 arteries per animal). Significant differences from controls were analyzed using unpaired t-test * P <0.05; ** P <0.01 vs control (Fig 3 A); or one way ANOVA followed by a Bonferroni test * P <0.05 and ** P <0.01 vs control; † P< 0.05 vs –E; # P <0.05 vs L-NOARG (Fig 3 B–E).

Article Snippet: Protein expression was quantified using primary antibodies anti-SK3 (Alomone, Israel, 1∶200 dilution), anti-IK1 (Alomone, Israel, 1∶450 dilution), or anti-β-actin as a loading control (Sigma-Aldrich, Spain, 1∶10000 dilution) and horseradish peroxidase conjugated secondary goat anti-mouse and anti-rabbit antibodies (Santa Cruz Biotech, CA, USA, 1∶10000 dilution).

Techniques: Inhibition

Journal: PLoS ONE

Article Title: Upregulation of SK3 and IK1 Channels Contributes to the Enhanced Endothelial Calcium Signaling and the Preserved Coronary Relaxation in Obese Zucker Rats

doi: 10.1371/journal.pone.0109432

Figure Lengend Snippet: (A, B) Immunohistochemical labeling of SK3 (A) and IK1 (B) channel in coronary arteries from LZR (top pannel) and OZR (bottom pannel). SK3 (Aa, Ac) and IK1 (Ba, Bc) channel immunoreactivity (arrows) was mainly located in the endothelium and eventually in the smooth muscle layer (asterisk) and was higher in arteries from OZR compared with LZR (Ac and Bc, respectively). Sections are representative of n = 3 OZR and n = 3 LZR, (1 artery per animal). (C, D) SK3 and IK1 channel expression is increased in coronary arteries from OZR. Western blot analysis of SK3 (C) and IK1 (D) channel expression in coronary arteries from LZR and OZR. Results were quantified by densitometry. Data are shown as means ± SEM of 5–7 animals. Significant differences from controls were analyzed using unpaired t-test * P <0.05; ** P <0.01 vs LZR.

Article Snippet: Protein expression was quantified using primary antibodies anti-SK3 (Alomone, Israel, 1∶200 dilution), anti-IK1 (Alomone, Israel, 1∶450 dilution), or anti-β-actin as a loading control (Sigma-Aldrich, Spain, 1∶10000 dilution) and horseradish peroxidase conjugated secondary goat anti-mouse and anti-rabbit antibodies (Santa Cruz Biotech, CA, USA, 1∶10000 dilution).

Techniques: Immunohistochemical staining, Labeling, Expressing, Western Blot

Journal: PLoS ONE

Article Title: Emerging Role of the Calcium-Activated, Small Conductance, SK3 K + Channel in Distal Tubule Function: Regulation by TRPV4

doi: 10.1371/journal.pone.0095149

Figure Lengend Snippet: A. RT-PCR analysis using whole kidney mRNA extracts revealed prominent bands of the appropriate size on agarose gels for both SK3 (473 bp) and BKα (318 bp), demonstrating expression of both of these channels in the kidney. SK3 primers were selected to cross the exon 2 and exon 3 borders to rule out amplification of intron sequences from genomic DNA. The electropherogram for SK3 is shown with both nucleotide sequences (NT) and amino acid sequences (AA) indicated for the segment across the exon border region, demonstrating that the PCR product does not originate from genomic DNA. 100-bp marker standards are shown (Lane M). B. Western blot of WT mouse kidney-SK3. SK3 protein is expressed as a single band near 90 kD in mouse kidney. SK3 blocking peptide (SK3-BP) was used as a control to verify antibody specificity which, as shown, abolished binding of the anti-SK3 antibody (right lane). Alpha-tubulin expression was used as a loading control (lower panel).

Article Snippet: Membranes were incubated with a well-characterized, high specificity, anti-SK3 primary antibody (anti-K Ca 2.3 directed against the N-terminus, 1∶100, Alomone Cat. #APC-025; see references – and Alomone web site) overnight at 4°C.

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Amplification, Marker, Western Blot, Blocking Assay, Binding Assay

Journal: PLoS ONE

Article Title: Emerging Role of the Calcium-Activated, Small Conductance, SK3 K + Channel in Distal Tubule Function: Regulation by TRPV4

doi: 10.1371/journal.pone.0095149

Figure Lengend Snippet: Antibodies and markers used for immunohistochemistry.

Article Snippet: Membranes were incubated with a well-characterized, high specificity, anti-SK3 primary antibody (anti-K Ca 2.3 directed against the N-terminus, 1∶100, Alomone Cat. #APC-025; see references – and Alomone web site) overnight at 4°C.

Techniques: Immunohistochemistry, Plasmid Preparation

Journal: PLoS ONE

Article Title: Emerging Role of the Calcium-Activated, Small Conductance, SK3 K + Channel in Distal Tubule Function: Regulation by TRPV4

doi: 10.1371/journal.pone.0095149

Figure Lengend Snippet: Top Panel (A–C): A low-magnification transverse section (5 µm) of the mouse kidney is shown. Discrete labeling is shown for staining for aquaporin-2 ( A. AQP2, red), a marker of the collecting ducts, SK3 ( B. SK3, green), and a merger of both channels ( C. Merge, yellow-organge for co-localization of AQP2 and SK3). Labeling is apparent for SK3 in both the cortex (label C) and medullary (label M) (dashed line shows cortical-medullary demarcation). Middle Pannel (D–F): Magnified view of the yellow inset box from A. SK3 co-localizes with all AQP2-postive tubules as show by the yellow-orange images (F., asterisk). SK3 staining is also apparent in AQP2-negative structures including other tubular structures (F., arrows) and smaller secondary structures (possibly vascular structures, F., arrow heads). Bottom Panel (G–H): Magnified view of staining in the presence of SK3 blocking peptide. All SK3 staining is abolished demonstrating specificity of our anti-SK3 antibody. Scale bar is 50 µm.

Article Snippet: Membranes were incubated with a well-characterized, high specificity, anti-SK3 primary antibody (anti-K Ca 2.3 directed against the N-terminus, 1∶100, Alomone Cat. #APC-025; see references – and Alomone web site) overnight at 4°C.

Techniques: Labeling, Staining, Marker, Blocking Assay

Journal: PLoS ONE

Article Title: Emerging Role of the Calcium-Activated, Small Conductance, SK3 K + Channel in Distal Tubule Function: Regulation by TRPV4

doi: 10.1371/journal.pone.0095149

Figure Lengend Snippet: Sagital section (5 µm) of WT mouse kidney showing staining for Tamm-Horsefall protein (THP, red), a marker of TAL cells, and SK3 (green). Panels A, C, E, and G are low magnification images showing THP staining of TAL structrues ( A ), SK3 labeling of the same structures (C), and a merged image (E). As shown at higher resolution for one of the tubules (inset from A), THP strongly stains the luminal border of the TAL ( B and F ) with SK3 also showing strong labeling of the luminal border and, to a variable degree, the abluminal border ( D and F ). The merged image ( F ) clearly identifies SK3 staining in the TAL cells. Panel H is a magnified view of a proximal tubule (PT), located left of the TAL in A . The PT showed minimal staining for SK3, although light staining was apparent along the luminal brush border. Scale bar is 10 µm.

Article Snippet: Membranes were incubated with a well-characterized, high specificity, anti-SK3 primary antibody (anti-K Ca 2.3 directed against the N-terminus, 1∶100, Alomone Cat. #APC-025; see references – and Alomone web site) overnight at 4°C.

Techniques: Staining, Marker, Labeling

Journal: PLoS ONE

Article Title: Emerging Role of the Calcium-Activated, Small Conductance, SK3 K + Channel in Distal Tubule Function: Regulation by TRPV4

doi: 10.1371/journal.pone.0095149

Figure Lengend Snippet: Mouse (WT) kidney section (5 µm) showing staining for the sodium-calcium exchanger (NCX, red), a marker of DCT, especially the later portion (DCT2), and SK3 (green). The heavy NCX staining of the upper portion of the tubule in Panel A (within yellow inset box) is consistent with the DCT2 segment with the weaker, more basolateral staining in the lower half of the tubule indicating this is the connecting tubule (CNT) (see text for details). Higher resolution image of the DCT2 ( D ) shows strong staining of SK3 along the luminal border with more variable, weaker staining along the abluminal border. The merged image clearly identifies SK3 staining of the DCT ( F ). In the CNT segment ( A. , labeled CNT), SK3 staining was also apparent along the luminal border with abluminal staining appearing weaker. Scale bar is 10 µm.

Article Snippet: Membranes were incubated with a well-characterized, high specificity, anti-SK3 primary antibody (anti-K Ca 2.3 directed against the N-terminus, 1∶100, Alomone Cat. #APC-025; see references – and Alomone web site) overnight at 4°C.

Techniques: Staining, Marker, Labeling

Journal: PLoS ONE

Article Title: Emerging Role of the Calcium-Activated, Small Conductance, SK3 K + Channel in Distal Tubule Function: Regulation by TRPV4

doi: 10.1371/journal.pone.0095149

Figure Lengend Snippet: Section (5 µm) from WT mouse kidney showing staining for AQP2 (red), a marker of PCs in collecting duct, and SK3 (green). Panels A, C, and E are low magnification views of a cross-section through a CCD identified by AQP2 staining. Panels B, D, and F represent a magnified view of the inset area from A (yellow inset box). Panel B shows strong AQP2 staining along the luminal border of PCs (5–6 cells), but not of the ICs (2 cells without staining). As shown in D and F , strong staining of SK3 is evident along the luminal border of all cells, both PCs and ICs. Variable, but weak staining, is also apparent along the abluminal border of some cells. However, the staining is most pronounced along the luminal border for both PCs and ICs, although typically stronger in PCs, as indicated by the SK3 fluorescence line intensity profiles across (luminal to abluminal direction) two cells identified as PC and IC ( Panel G ). H . Relative mean intensity profiles (± SEM) across the cells from all sections showing the maximal values across the luminal border (Apical) and abluminal border (Basal) and the minimal values within the cytoplasm (Cytosol). The mean values are given for both PCs (n = 37) and ICs (n = 12) from all sections analyzed. The maximal luminal intensity is much greater than the abluminal intensity (*P<0.02) indicating dominant expression at the luminal border. Scale bar is 10 µm.

Article Snippet: Membranes were incubated with a well-characterized, high specificity, anti-SK3 primary antibody (anti-K Ca 2.3 directed against the N-terminus, 1∶100, Alomone Cat. #APC-025; see references – and Alomone web site) overnight at 4°C.

Techniques: Staining, Marker, Fluorescence, Expressing

Journal: PLoS ONE

Article Title: Emerging Role of the Calcium-Activated, Small Conductance, SK3 K + Channel in Distal Tubule Function: Regulation by TRPV4

doi: 10.1371/journal.pone.0095149

Figure Lengend Snippet: A. Fluorescence image of a split-open CCD loaded with the voltage-sensitive fluorescence dye, DiSBAC 2 (3), showing loading of all cells. The fluorescence intensity is an index of Vm and is presented as relative fluorescence units (RFU). B. Effect of 50 mM K + (High K + ) application on Vm of CCD cells showing the expected membrane depolarization (increased RFU). C. Effect of 300 nM apamin or 50 nM IbTX application on Vm in basal conditions showing little or no effect of either apamin (Apa) or IbTX in the basal state (TRPV4 not activated). D. Effect of TRPV4 activation with GSK101 (50 nM) leading to membrane hyperpolarization of Vm (decreased RFU), as expected for SK3 and BK activation. Subsequent application of either 300 nM apamin or 50 nM IbTX now induce a marked depolarization of Vm (increased RFU) demonstrating inhibition of SK3 and BK, respectively. E. Summary graph showing mean changes in Vm in basal conditions upon addition of High K + (High K + , n = 44 cells), 300 nM apamin, or 50 nM IbTX (Left panel, Basal). Right panel (GSK101: TRPV4 Activation) shows the results after activation of TRPV4 (Ca 2+ influx). Both apamin and IbTX now bring about a significant depolarization of Vm (*P<0.01 compared to Basal). The combine addition of both apamin and IbTX (Apa + IbTX) displays an enhanced depolarization compared to addition of apamin or IbTX alone (**P<0.01). The number in parentheses is the number of cells for each group (n).

Article Snippet: Membranes were incubated with a well-characterized, high specificity, anti-SK3 primary antibody (anti-K Ca 2.3 directed against the N-terminus, 1∶100, Alomone Cat. #APC-025; see references – and Alomone web site) overnight at 4°C.

Techniques: Fluorescence, Activation Assay, Inhibition

Journal: Cerebral cortex (New York, N.Y. : 1991)

Article Title: Apamin Improves Prefrontal Nicotinic Impairment in Mouse Model of Alzheimer's Disease.

doi: 10.1093/cercor/bhz107

Figure Lengend Snippet: Figure 3. There are no significant genotype differences in Fluo-5F calcium responses elicited by nicotinic stimulation, nor in the protein levels for SK2 and SK3 channels in the PFC. In the presence of the muscarinic receptor antagonist atropine, bath application of acetylcholine elicited large Ca2+ responses in layer 6 pyramidal cells of PFC in both WT and TgCRND8 animals. Representative examples of quantified Fluo-5F Ca2+ fluorescence and accompanying images are shown for a (A) WT neuron and a (B) TgCRND8 neuron. Scale bar, 5 μm. Graphs show (C) peak Ca2+ response and (D) area under the curve (AUC) of the dF/F traces. (E–G) Furthermore, there are no significant differences in SK2 and SK3 channel levels in the PFC of WT and TgCRND8 mice. (E) Example immunoblot images for SK2 and SK3 shown with stain-free total membrane protein used as a loading control for band normalization, with alternating samples from WT control mice (blue) and TgCRND8 mice (red). Graphs show (F) SK2 and (G) SK3 band intensity for WT and TgCRND8 mice normalized against total membrane protein, with results from each mouse represented as a percent of the respective WT mean.

Article Snippet: The Frontier Institute SK2 and the Alomone SK3 primary antibodies were validated in SK2 knockout by immunohistochemistry (Lin et al., 2008, in its supplemental section) and SK3 knockout by western blot (Bond et al., 2004), respectively.

Techniques: Fluorescence, Western Blot, Staining, Membrane, Control

Journal: Biochimica et biophysica acta

Article Title: Identity and function of a cardiac mitochondrial small conductance Ca 2+ -activated K + channel splice variant

doi: 10.1016/j.bbabio.2017.03.005

Figure Lengend Snippet: Specific primers for three SKCa isoforms (SK1-3) amplification from mRNA of isolated guinea pig ventricular myocyte.

Article Snippet: One membrane was incubated with specific primary antibody anti-SK3 (Alomone Lab APC-103) (SK3 left panel); another membrane was incubated with the same antibody, anti-SK3, but which was neutralized with the corresponding immunizing SK3 peptide before being added (SK3 preincubation, right panel).

Techniques: Amplification, Isolation

Journal: Biochimica et biophysica acta

Article Title: Identity and function of a cardiac mitochondrial small conductance Ca 2+ -activated K + channel splice variant

doi: 10.1016/j.bbabio.2017.03.005

Figure Lengend Snippet: Expression of mSKCa channels in guinea pig hearts. A: A representative agarose gel analysis of RT-PCR amplified SK1, SK2 and SK3 from guinea pig cardiomyocytes (lanes 1–6) and human ventricular tissue (lanes 9–10). Lane 1: SK1-N; Lane 2: SK1-C; Lane 3: SK2-N; Lane 4: SK2-C; Lane 5: SK3-N; Lane 6: SK3-C; Lane 9: human SK3-N; Lane 10: human SK3-C; lanes 7 and 8: DNA ladder; Lane 11: RNA control. B: Alignment of SK3 splice variants. Red: deduced amino acid (a.a.) sequences from amplified genes; Blue: P-W sequence designating SK3.2; Black dashes: guinea SK3 transmembrane (T) domains 1–6; CaMBD: Ca2+ calmodulin binding domain; Bold: human vs. guinea pig a.a. differences; Yellow: truncated N terminus (a.a. 1–277) or C-terminus (a.a. 627–720) of SK3.1 as overexpressed in HL-1 cells (Fig. 8). C: Western blot analyses of SK3 in guinea pig heart (Heart), heart mitochondria (Mito) and mitochondrial inner membrane (IMM). One membrane was incubated with specific primary antibody anti-SK3 (Alomone Lab APC-103) (SK3 left panel); another membrane was incubated with the same antibody, anti-SK3, but which was neutralized with the corresponding immunizing SK3 peptide before being added (SK3 preincubation, right panel). Tubulin, GM130, and calnexin were used to monitor purity of mitochondria and IMM; no contamination by cytosol, Golgi, or sarcoplasmic reticular membrane protein was observed in mitochondria and IMM. ANT was used to monitor integrity of mitochondria and IMM. Since SK3 expression is weak in mitochondria, the protein concentration loaded in the mito lane was larger than that in heart and IMM lanes.

Article Snippet: One membrane was incubated with specific primary antibody anti-SK3 (Alomone Lab APC-103) (SK3 left panel); another membrane was incubated with the same antibody, anti-SK3, but which was neutralized with the corresponding immunizing SK3 peptide before being added (SK3 preincubation, right panel).

Techniques: Expressing, Agarose Gel Electrophoresis, Reverse Transcription Polymerase Chain Reaction, Amplification, Sequencing, Binding Assay, Western Blot, Incubation, Protein Concentration

Journal: Biochimica et biophysica acta

Article Title: Identity and function of a cardiac mitochondrial small conductance Ca 2+ -activated K + channel splice variant

doi: 10.1016/j.bbabio.2017.03.005

Figure Lengend Snippet: Representative recordings of mK+ uptake activated by adding CaCl2 to buffer in HL-1 cells transfected with SK3.1 constructs. Traces of PBFI fluorescence indicate changes in m [K+]. A: in SK3FL-EGFP transfected HL-1 cells; B: in SK3FL-EGFP transfected HL-1 cells with alamethicin, a mitochondrial pore forming peptide; C: in pEGFP-N3 transfected HL-1 cells; D: in SK3Δ626–720–EGFP transfected HL-1 cells; and E: in SK3FL-EGFP transfected HL-1 cells with apamin, a blocker of the SK3 channel. F: averaged ΔPBFI ratios induced by adding CaCl2 to permeabilized HL-1 cells. n= 20 cells; *P < 0.05 vs. EGFP control. Because of the presence of 3 mM EGTA and 5 mM MgCl2, mitochondrial [Ca2+] with added 2 mM CaCl2 remained within the nM range.

Article Snippet: One membrane was incubated with specific primary antibody anti-SK3 (Alomone Lab APC-103) (SK3 left panel); another membrane was incubated with the same antibody, anti-SK3, but which was neutralized with the corresponding immunizing SK3 peptide before being added (SK3 preincubation, right panel).

Techniques: Transfection, Construct, Fluorescence

Journal: Biochimica et biophysica acta

Article Title: Identity and function of a cardiac mitochondrial small conductance Ca 2+ -activated K + channel splice variant

doi: 10.1016/j.bbabio.2017.03.005

Figure Lengend Snippet: Location of mSK3 in human heart and rat cardiomyocytes. A: Immuno-electron microscopic (IEM) detection of isolated human ventricular mitochondria immunostained with specific antibody against SK3. B: Western blot analyses of SK3 expression in human ventricular tissue (heart), mitochondria (mito) and inner mitochondrial membrane (IMM). Tubulin confirmed purity of mitochondrial isolation, adenine nucleotide translocase (ANT) confirmed enrichment of mitochondria and IMM, and GM130 and calnexin confirmed absence of contamination by Golgi or sarcoplasmic reticulum membrane, respectively. C: Confocal fluorescence images of adult rat cardiomyocytes immuno-stained simultaneously with SK3 antibody followed by FITC conjugated secondary antibody (middle panel, Green) and COX 1 (cytochrome c oxidase) antibody, followed by Alexa Fluor 546 conjugated secondary antibody (top panel, red). Yellow color in merged image (bottom panel) depicts SK3 localized in mitochondria; the intensity profile panel showed the overlapping degree of SK3 with COX 1. The x-axis in the intensity profile is from top to bottom. The yellow rectangles label the overlap areas of SK3 with COX 1.

Article Snippet: One membrane was incubated with specific primary antibody anti-SK3 (Alomone Lab APC-103) (SK3 left panel); another membrane was incubated with the same antibody, anti-SK3, but which was neutralized with the corresponding immunizing SK3 peptide before being added (SK3 preincubation, right panel).

Techniques: Isolation, Western Blot, Expressing, Fluorescence, Staining

Journal: Biochimica et biophysica acta

Article Title: Identity and function of a cardiac mitochondrial small conductance Ca 2+ -activated K + channel splice variant

doi: 10.1016/j.bbabio.2017.03.005

Figure Lengend Snippet: SK3.1 localized into mitochondria. Top panels: Schematic of SK3FL-EGFP, SK3Δ1–277–EGFP and SK3Δ626–720–EGFP constructs. Q sequence, poly glycine sequence; T1–6, transmembrane regions; P, selectivity pore; CaMBD, Ca2+ calmodulin binding domain; EGFP, enhanced green fluorescence protein. Bottom panels: Confocal fluorescence images of HL-1 cells transfected with SK3FL-EGFP (A, Green), SK3Δ626–720–EGFP (B, Green) or SK3Δ1–277–EGFP (C, Green) and stained with Mitotracker red, (MTR, Red). Merging (orange) depicts colocalization of SK3 with MTR; the intensity profiles show the overlapping degree of SK3 with MTR. The x-axis in the intensity profile was recorded from top to bottom and from left to right for regions 1 and 2 respectively. The yellow rectangles label the overlap areas of SK3 with MTR.

Article Snippet: One membrane was incubated with specific primary antibody anti-SK3 (Alomone Lab APC-103) (SK3 left panel); another membrane was incubated with the same antibody, anti-SK3, but which was neutralized with the corresponding immunizing SK3 peptide before being added (SK3 preincubation, right panel).

Techniques: Construct, Sequencing, Binding Assay, Fluorescence, Transfection, Staining

Journal: Biochimica et biophysica acta

Article Title: Identity and function of a cardiac mitochondrial small conductance Ca 2+ -activated K + channel splice variant

doi: 10.1016/j.bbabio.2017.03.005

Figure Lengend Snippet: Lactate dehydrogenase (LDH) release is reduced if SK3 C-terminus is intact. HL-1 cells transfected with pEGFP-N3, SK3FL, SK3Δ1–277 and SK3Δ626–720 were subjected to hypoxia/reoxygenation or normoxia. LDH activity was measured in the reoxygenation medium. Data presented are means ± SE from 3 independent experiments. *P < 0.05 vs. normoxia groups, #P < 0.05 vs. EGFP groups.

Article Snippet: One membrane was incubated with specific primary antibody anti-SK3 (Alomone Lab APC-103) (SK3 left panel); another membrane was incubated with the same antibody, anti-SK3, but which was neutralized with the corresponding immunizing SK3 peptide before being added (SK3 preincubation, right panel).

Techniques: Transfection, Activity Assay

Journal: Biochimica et biophysica acta

Article Title: Identity and function of a cardiac mitochondrial small conductance Ca 2+ -activated K + channel splice variant

doi: 10.1016/j.bbabio.2017.03.005

Figure Lengend Snippet: Silencing SK3 in HL-1 cells increases cell apoptosis. A: Real-time PCR assessment of SK3 relative mRNA abundance in HL-1 cells transfected with SK3 silence siRNA or scramble siRNA. Data presented are means ± SE from 3 independent experiments. * P < 0.05 vs. scramble siRNA groups. B: Representative TUNEL staining of HL-1 cells transfected with SK3 siRNA or scramble siRNA and subjected to normoxia (control) or to 2 h hypoxia and 16 h reoxygenation. Apoptotic nuclei were TUNEL stained (red) and counterstained with DAPI (blue) to label nuclei. C: Quantitative analysis of TUNEL-positive HL-1 cells. Mean ± SE, n = 2000 HL-1 cells per group. * P < 0.05 vs. scramble siRNA, # P < 0.05 vs. normoxia.

Article Snippet: One membrane was incubated with specific primary antibody anti-SK3 (Alomone Lab APC-103) (SK3 left panel); another membrane was incubated with the same antibody, anti-SK3, but which was neutralized with the corresponding immunizing SK3 peptide before being added (SK3 preincubation, right panel).

Techniques: Real-time Polymerase Chain Reaction, Transfection, TUNEL Assay, Staining

Journal: Biochimica et biophysica acta

Article Title: Identity and function of a cardiac mitochondrial small conductance Ca 2+ -activated K + channel splice variant

doi: 10.1016/j.bbabio.2017.03.005

Figure Lengend Snippet: Silencing SK3 in HL-1 cells attenuates ΔΨm. HL-1 cells were transfected with SK3 silenced siRNA or scramble siRNA. Changes in TMRM fluorescence (ΔΨm) was measured in HL- 1 cells subjected to simulated ischemia (blocked respiration). A: Time course of relative changes in ΔΨm was assessed in SK3 siRNA (n = 40 cells) or in scramble siRNA (n =50 cells) transfected HL-1 cells exposed to 10 min simulated ischemia followed by 8 min resuffusion of cells with normal Tyrode to wash out the simulated ischemia buffer and restore control conditions. (Time control = no ischemia; n =45 cells). *P < 0.05 vs. scramble siRNA. B: Representative confocal images of TMRM fluorescence in HL-1 cells transfected with SK3 siRNA or scramble siRNA at conditions and times noted in A.

Article Snippet: One membrane was incubated with specific primary antibody anti-SK3 (Alomone Lab APC-103) (SK3 left panel); another membrane was incubated with the same antibody, anti-SK3, but which was neutralized with the corresponding immunizing SK3 peptide before being added (SK3 preincubation, right panel).

Techniques: Transfection, Fluorescence