MA3-925

antibody from Invitrogen Antibodies

Targeting: RYR1 CCO, MHS, MHS1, PPP1R137, RYR

Provider product page for MA3-925

Western blot

Immunohistochemistry

Other assay

Antibody data

Antibody Data
Antigen structure
References [199]
Comments [0]
Validations
Western blot [1]
Immunohistochemistry [1]
Other assay [13]

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Product number: MA3-925 - Provider product page
Provider: Invitrogen Antibodies
Product name: RyR1 Monoclonal Antibody (34C)
Antibody type: Monoclonal
Antigen: Purifed from natural sources
Description: MA3-925 detects ryanodine receptor (RyR)-1 and RyR-2 isoforms in amphibian, chicken, bovine, canine, fish, human, non-human primate, mouse, rabbit, mink, rat and sheep tissues. In chickens this antibody detects the alpha, beta and cardiac isoforms. This antibody also detects RyR-3 in mouse cells. In frog, MA3-925 detects the alpha and beta isoforms and in fish it detects the alpha isoform.
Antibody clone number: 34C
Concentration: 1 mg/mL

RYR1 protein structure - MA3-925 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 3 Ca 2+ handling proteins. (A) Representative western blot images of Ca V 1.1, RyR1, SERCA1, SERCA2, calsequestrin (1 and 2), and sarcolipin. To allow the use of data in multiple western blots each band within a single western blot was normalized to GAPDH for that sample as a loading control and then normalized to the average WT values for that particular gel to give %WT. (B) Analysis of muscle levels of Ca V 1.1 normalized to GAPDH. (C) Analysis of muscle levels of RyR1 normalized to GAPDH. (D) Analysis of muscle levels of SERCA1 normalized to GAPDH. (E) Analysis of muscle levels of SERCA2 normalized to GAPDH. (F) Analysis of muscle levels of CSQ normalized to GAPDH. (G) Analysis of sarcolipin normalized to GAPDH. (H) SERCA activity as a function of Ca 2+ concentration. (I) Scheme of changes in Ca 2+ handling proteins. Values are shown as mean +- SEM. * P

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 7 Fat1 expression at late stages of muscle differentiation. ( A ) Fat1 expression was visualized in E13.5 embryos or in neonate (P0) muscle by beta-galactosidase staining or by in situ hybridization with a Fat1 3'UTR RNA probe. ( B - D ) Immunolocalization of FAT1 (anti-FAT1-ICD from [35] , green) was performed in E12.5 mouse embryo ( C ), and on adult ( B, D ) muscle fibers on longitudinal muscle cryosections from wild type ( B , C 1-3 , D 1,4 ), from Fat1 DeltaTM/DeltaTM embryos ( C 4-6 ), and from Fat1 LacZ/LacZ ( D 2-3 , D 5-6 ) mice, combined with either antibodies against alpha-actinin (red, B 5 ), DHPR (Cacna1s) (red, B 2,3 ), or RyR (red, B 4 ), or with Phalloidin (red, C, D ). In D , Green channel images (FAT1) were first captured with either identical exposure time between wild type and mutants ( D 1,4 and D 2,5 , 421 ms), or with longer exposure time ( D 3,6 , 2222 ms). This indicates that the epitope detected by the anti-FAT1-ICD antibody (from ref [35] ) is present in reduced but detectable amounts in Fat1 LacZ/LacZ muscles. This observation was made when Fat1 LacZ/LacZ mice (n = 2 at P0; and n = 3 at adult stages) displayed severe muscle defects at the stage of dissection, indicating that levels of FAT1 protein inversely correlate with phenotype severity. Scale bars: ( B - D ) 4 um, ( C ) 6 um.

Supportive validation

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Experimental details: Fig 2 PLN phosphorylation and expression levels of other Ca 2+ regulatory proteins. (A) Western blot analyses demonstrating that monomeric p-PLN/PLN ratio is significantly lower in both Pln OE and Pln OE / Sln KO soleus muscles compared with WT (n = 5-6 per genotype). Western blotting for other major Ca 2+ regulatory proteins SERCA1a (B, 110 kDa), SERCA2a (C, 110 kDa), RyR (D, 565 kDa), DHPR (E, 170 kDa), and CSQ (F, 63 kDa) did not reveal any significant differences across genotypes (n = 5-8 per genotype, with a 6 mug load for all proteins). *Significantly different from WT using a a one-way ANOVA with a Tukey's post-hoc, p

Supportive validation

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Experimental details: Fig. 1 Actomyosin ATPase and sarcoplasmic reticulum (SR) Ca 2+ -ATPase activities are decreased in AIA EDL muscles. Representative Coomassie brilliant blue staining of myofibrillar proteins ( a ), percentage distribution of myosin heavy chain (MyHC) and actin content in total myofibrillar proteins ( b ), and actomyosin ATPase activity ( c ). Electrophoretically separated MyHC isoforms ( d ) and percentage distribution of MyHC isoforms ( e ): I, slow isoform and IIa, IId/x, and IIb, fast isoforms. Representative Western blots of ryanodine receptor (RyR1), dihydropyridine receptor alpha2 subunit (DHPR), and SR Ca 2+ -ATPase (SERCA) 1 ( f ), the expression levels of these proteins normalized to actin content ( g ), and SERCA activity ( h ). Control (CNT), white bars ; AIA, black bars . Data are presented as mean +- SEM for three to seven muscles in each group. * P < 0.05 versus CNT

Supportive validation

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Experimental details: Figure 1. Capn3 deficiency in mouse C2C12 myotubes reduces SERCA protein levels and SERCAfunction. (a) C2C12 myotubes treated with NS or Capn3 shRNAs and differentiated for7 days. Scale bar = 100 mum (b) Western blot analysis showing significant decreaseof Capn3 (SPA antibody), SERCA1, SERCA2 and RyR1 levels in Capn3 knockdown myotubes,compared with controls (* P < 0.05;** P < 0.01). Total levels of DHPR, TRPC1, MyHC and actin arenot significantly changed. N = 3 independent experiments run on thesame gel. (c) SERCA-specific ATPase activity determined in homogenates from C2C12myotubes. Capn3-deficient myotubes show a significant reduction of SERCA activitycompared with NS-controls ( n = 3,* P < 0.05). (d and e) Ca 2+ imaging of C2C12myotubes loaded with Fura2-AM shows delayed Ca 2+ clearance from thecytosol in Capn3 knockdown myotubes. (d) Two representative traces of changes inFura2-AM fluorescence ratios( F 340 / F 380 ) fromCapn3-shRNA and NS-shRNA treated myotubes. Ca 2+ transients were elicitedby local stimulation with KCl 50 m m in the absence of extracellularCa 2+ . (e) Tau, the time constant of the Ca 2+ transient decayphase in seconds (s), is significantly increased in Capn3-deficient myotubes.** P < 0.01, n = total number of myotubesrecorded from six different experiments are shown in the graph.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2. CAPN3 deficiency in human myotubes reduces SERCA protein levels and SERCA function.(a) LHCN-M2 myoblasts treated with control NS-shRNA or CAPN3-shRNAs anddifferentiated for 9 days. Scale bar = 50 mum. (b) Representative Western blotanalysis showing decrease of CAPN3 (12A2 mAb), SERCA1, SERCA2 and RyR1 proteinlevels in CAPN3-sh treated myotubes compared with controls. DHPRalpha2, MyHC and actinlevels remain unaltered. (c) SERCA-specific ATPase activity determined inhomogenates from LHCN-M2 myotubes. CAPN3-deficient myotubes show a significantreduction of SERCA activity compared with NS-controls ( n = 3,* P < 0.05). D-F) Ca 2+ imaging of LHCN-M2myotubes loaded with Fura2-AM shows increased cytosolic [Ca 2+ ] anddelayed Ca 2+ clearance from the cytosol in CAPN3 knockdown myotubes. (d)Resting cytosolic [Ca 2+ ] was measured in the presence of 2 m m Ca 2+ at 37degC. CAPN3-deficient human myotubes show significantlyincreased resting cytosolic [Ca 2+ ] compared with controls(* P < 0.05; n = 10 experiments). Totalnumbers of myotubes recorded are shown in the graph. (e) Two representative tracesof changes in Fura2-AM fluorescence ratios( F 340 / F 380 ) fromCAPN3-shRNA and NS-shRNA treated myotubes. Ca 2+ transients were elicitedby local stimulation with KCl 130 m m in the absence of extracellularCa 2+ . (f) Tau, the time constant of the Ca 2+ transient decayphase in seconds (s), is significantly increased in CAPN3-deficient myotubes.* P < 0.05, n = total number of myotubesr

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 5 Expression level of skeletal muscle proteins, the amount of releasable Ca 2+ from the SR to the cytosol, and the resting cytosolic Ca 2+ level in primary skeletal myotubes. ( a ) The immunoblot analysis of proteins mediating Ca 2+ movements and handling in skeletal muscle was conducted using the lysate of myotubes treated with sildenafil. Fifteen proteins were examined, and no expression levels were changed by sildenafil (the expression level of proteins is presented as bar graphs in Supplementary Figure 5 ). alpha-Actin was used as a loading control. At least three independent experiments per protein were conducted and a representative result is presented. JP, junctophilin; CSQ, calsequestrin; Mg, mitsugumin; CaM, calmodulin. The amount of releasable Ca 2+ from the SR to the cytosol in response to TG (2.5 mu M ) ( b ) or resting cytosolic Ca 2+ level ( c ) was examined in myotubes treated with sildenafil, and histograms are shown for the normalized peak amplitude or area under the peak to the mean value of those from untreated controls as described in the 'Materials and methods' section. TG was applied to myotubes in the absence of extracellular Ca 2+ to avoid extracellular Ca 2+ entry. The results are presented as the means+-s.e. for the number of experiments presented in the parentheses of Table 2 . *, and the significant difference was compared with the untreated control ( P

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 3 Effect of the IT mutation on triadin and CSQ. ( a ) Representative western blots of triadin levels in soleus muscle from WT and IT mice at different ages. ( b ) Analysis of triadin levels. ( c ) RyR1 and CSQ localization in FDB fibres by immunocytochemistry. Magenta: RyR1, Green: CSQ. ( d , e ) Line scans of CSQ and RyR1 immunostaining in FDB fibres of IT ( e ) and WT ( d ) mice. ( f ) Analysis of the fluorescence profile of CSQ in WT and IT fibres, FWHM is full width at half maximum. Data are shown as mean+-s.e.m. * P

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig 2 Neither contractile proteins nor excitation-contraction coupling proteins are altered in PH diaphragm muscles. A : Stain free images of myosin heavy chain (MyHC) and western blots of troponin (Tn) T. B : the expression levels of MyHC or TnT normalized to actin content. C : electrophoretically separated MyHC isoforms. D : percentage distribution of MyHC isoforms: I, slow myosin isoform; IIa, IId/x, and IIb, fast myosin isoforms. E : representative western blots of ryanodine receptor (RyR1), dihydropyridine receptor alpha2 subunit (DHPR), sarcoplasmic reticulum Ca 2+ -ATPase (SERCA) 1, and SERCA2. F : the expression levels of RyR1, DHPR, SERCA1, or SERCA2 normalized to actin content. Control (CNT), white bars; AIA, black bars. Data represent mean +- SD for 4-8 rats in each group.