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Gamma CA | Gamma Carbonic anhydrases

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AS17 4114  |  Clonality: Polyclonal  |  Host: Rabbit  |  Reactivity: Arabidopsis thaliana

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AS17 4114

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product information
Background

Gamma CAH (Gamma Carbonic anhydrase) proteins are localized in the inner mitochondrial membrane. They are in a range of 25-30 kDa.

Immunogen

KLH-conjugated peptide derived from Arabidopsis thaliana Gamma-Carbonic anhydrase, UniProt: Q9C6B3, TAIR:  AT1G47260

Host Rabbit
Clonality Polyclonal
Clone
Purity Serum
Format Lyophilized
Quantity 50 µl
Reconstitution For reconstitution add 50 ĩl of sterile water. Serum contains 0.1 % sodium azide as preservative.
Storage Store lyophilized/reconstituted at -20°C; once reconstituted make aliquots to avoid repeated freeze-thaw cycles. Please, remember to spin tubes briefly prior to opening them to avoid any losses that might occur from lyophilized material adhering to the cap or sides of the tubes.
Tested applications Western blot (WB)
Related products Mitochondrial antibody collection.

Secondary antibodies

Additional information
application information
Recommended dilution 1 : 1000 (WB)
Expected | apparent MW 9.97 KDa (gamma-CA1), 30KDa (gamma-CA2) strongest band, 27.83 (gamma-CA3), 25.08 KDa Arabidopsis thaliana proteins
Confirmed reactivity Arabidopsis thaliana
Predicted reactivity Higher plants
Not reactive in
Additional information
Selected references Kühn et al. (2015). Complete Mitochondrial Complex I Deficiency Induces an Up-Regulation of Respiratory Fluxes That Is Abolished by Traces of Functional Complex I. Plant Physiol. 2015 Aug;168(4):1537-49. doi: 10.1104/pp.15.00589. Epub 2015 Jul 1.

application example

western blot using anti-plant gamma carbonic anhydrase

Mitochondrial proteins (1mg) from Arabidopsis thaliana ecotype Col-0 cell suspensions were separated by 2D BN/SDS-PAGE (Klodmann et al., 2011; Plant Physiology 157: 587-598). After gel electrophoresis, proteins were transferred to nitrocellulose membrane using semi-dry conditions (90 min; 0.8mA per cm2 of gel). After washing once in TBS-T buffer (10 min), blot was blocked overnight at 4ºC with 10% p/v milk in TBS-T. Blot was washed three times with 0.5% p/v milk in TBS-T (10 min each) and incubated 1 hour with primary antibody (1:1000 in TBS-T, 3% BSA and 0.02% NaN3). Washing was carried out three times with TBS-T, 0.5% BSA, 10 min each. Then blot was incubated 1 hour with secondary antibody (Anti-rabbit IgG conjugated to alkaline phosphatase at a dilution of 1:10 000) in TBS-T, 3% BSA, 0.02% NaN3). After washing (as above), blot was equilibrated 5 minutes in AP buffer (Tris-HCl 1M pH 9.5; 5M NaCl; 4M MgCl2). Finally, revealing was developed by incubation in AP buffer; 110 µg/ml NBT; 75 µg/ml BCiP. Reaction was stopped by discarding revealing solution and adding distilled water. Upper lane: 1D BN gel electrophoresis of mitochondrial proteins, with respective complexes.

Lower gel: second dimension, where mitochondrial proteins of complexes were separated. Blot section (indicated as dashed square in gel) reveals mainly two bands (around 30 kDa- arrows), corresponding to both gamma-CA2 forms. The smear along the 30 kDa line is interpreted as fragments of complex I containing CA2 protein (Perales et al., 2005; Journal of Molecular Biology, 350: 263-277).

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