rabbit polyclonal antibody against ca v 1 2  (Alomone Labs)

Journal: Translational Psychiatry

Article Title: Identification of ultra-rare disruptive variants in voltage-gated calcium channel-encoding genes in Japanese samples of schizophrenia and autism spectrum disorder

doi: 10.1038/s41398-022-01851-y

Figure Lengend Snippet: a Three-dimensional (3D) model structure of the Ca v 1.2 channel with the folded N-terminal structure. b 3D model structure of the Ca v 1.2 channel with the CaM-binding structure, shown in complex with the N-lobe of CaM. c Enlarged view around the alanine 36 (A36) residue of the folded N-terminal structure. The A36 site is highlighted by green dotted circles. The N-terminal spatial Ca 2+ -transforming element (NSCaTE) region (47–68), Ca 2+ , and CaM are indicated in yellow, orange, and magenta, respectively. Molecular graphics were created using UCSF Chimera . d A schematic illustration of the hypothesis that the A36V mutation attenuates Ca 2+ -dependent inactivation (CDI) by conformational equilibrium shift favoring the folded structure.

Article Snippet: The antibodies used for immunoblotting were purchased commercially as follows: a rabbit polyclonal antibody against Ca v 1.2 (ACC-003, Alomone Labs, Jerusalem, Israel), a mouse monoclonal antibody against β-actin (A5441, Merck), and secondary antibodies conjugated to HRP (ab97051 and ab97023, abcam, Cambridge, UK).

Techniques: Binding Assay, Mutagenesis

Journal: Translational Psychiatry

Article Title: Identification of ultra-rare disruptive variants in voltage-gated calcium channel-encoding genes in Japanese samples of schizophrenia and autism spectrum disorder

doi: 10.1038/s41398-022-01851-y

Figure Lengend Snippet: a Sanger sequencing results for the de novo variant p.A36V (left) and schematic illustration of the primary structure of the Ca v 1.2 channel (right). The red asterisk indicates the A36V mutation near the N-terminal spatial Ca 2+ -transforming element (NSCaTE). b The N-terminal amino acid sequences for Ca v 1.2 channels (short isoforms). The A36V mutation and the A39V Brugada mutation are indicated in red letters. c Expression of wild-type (WT) and A36V Ca v 1.2 channels in HEK293T cells as detected by anti-Ca v 1.2 antibody. d Membrane localization of WT and A36V Ca v 1.2 channels overexpressed in BHK cells. The plasma membrane was visualized by membrane-tethering red fluorescent protein (RFP-KRasCT). e The fluorescence intensity profiles of the line shown in Fig. 1d. f Plasma membrane to cytoplasm intensity ratio of Ca v 1.2. Statistical comparison was performed by two-tailed Welch’s t test (n.s., not significant). Data are presented as mean ± s.e.m.

Article Snippet: The antibodies used for immunoblotting were purchased commercially as follows: a rabbit polyclonal antibody against Ca v 1.2 (ACC-003, Alomone Labs, Jerusalem, Israel), a mouse monoclonal antibody against β-actin (A5441, Merck), and secondary antibodies conjugated to HRP (ab97051 and ab97023, abcam, Cambridge, UK).

Techniques: Sequencing, Variant Assay, Mutagenesis, Expressing, Fluorescence, Two Tailed Test

Journal: Translational Psychiatry

Article Title: Identification of ultra-rare disruptive variants in voltage-gated calcium channel-encoding genes in Japanese samples of schizophrenia and autism spectrum disorder

doi: 10.1038/s41398-022-01851-y

Figure Lengend Snippet: a Families of Ba 2+ currents evoked by 30-ms depolarizing pulses from −30 to 60 mV with increments of 10 mV for wild-type (WT) and A36V neuronal Ca v 1.2 channels. b Current density–voltage ( I – V ) relationships. Data are expressed as mean ± s.e.m., WT: n = 18, A36V: n = 12. The values of G, Erev, V 0.5 , and k were −0.40, 63.0 mV, 7.6 mV, and 5.6 mV for WT channels, and −0.50, 61.3 mV, 6.7 mV, and 4.9 mV for A36V Ca v 1.2 channels. c Inactivation curves for WT (○, n = 9) and A36V (●, n = 4) neuronal Ca v 1.2 channels. Data are expressed as mean ± s.e.m. The values of V 0.5 , and k were (respectively) −37.6 mV and 11.5 mV for WT channels, and −41.6 mV and 12.1 mV for A36V Ca v 1.2 channels. d , g Ca 2+ -dependent inactivation (CDI) of neuronal ( d ) and cardiac ( g ) Ca v 1.2 channels. Ba 2+ (blue) and Ca 2+ (black) currents evoked by 350-ms step depolarization to 30 mV were normalized at their peak current amplitudes for WT and A36V Ca v 1.2 channels. e , f, h, i , Ratios of current amplitude to the peak amplitude were plotted against depolarizing time in the Ba 2+ ( e, h ) and the Ca 2+ ( f, i ) external solutions. The numbers of recorded cells were 10 and 15 for WT and A36V neuronal Ca v 1.2 channels ( e , f ), and 8 and 6 for WT and A36V cardiac Ca v 1.2 channels ( h – i ), respectively. Statistical comparison was performed by two-tailed non-paired Student’s t test (* p < 0.05). Data are presented as mean ± s.e.m.

Article Snippet: The antibodies used for immunoblotting were purchased commercially as follows: a rabbit polyclonal antibody against Ca v 1.2 (ACC-003, Alomone Labs, Jerusalem, Israel), a mouse monoclonal antibody against β-actin (A5441, Merck), and secondary antibodies conjugated to HRP (ab97051 and ab97023, abcam, Cambridge, UK).

Techniques: Two Tailed Test