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Cyt c | Cytochrome c

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AS08 343A  |  Clonality: Polyclonal  |  Host: Rabbit  |  Reactivity: Arabidopsis thaliana, Lupinus luteus |  Marker of PCD

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AS08 343A

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product information
Background

Cytochrome c is located in inner mitochondrial membrane. It is a small heme protein which, unlike other cytochromes, is highly soluble. This protein is an essential component of the electron transport chain, where it undergoes oxidation and reduction without binding oxygen.

Immunogen

KLH-conjugated synthetic peptide derived from Arabidopsis thaliana cytochrome c protein sequence, UniProt:D7KMK0 (C-1) D7LY03 (C-2), TAIR: At1g22840 (Cytc1) and At4g10040 (Cytc2)

Host Rabbit
Clonality Polyclonal
Clone
Purity Affinity purified serum in PBS, pH 7.4. 
Format Lyophilized
Quantity 50 µg
Reconstitution
For reconstitution add 50 µl of sterile water.
Storage

Store lyophilized at -20°C; once reconstituted make aliquots to avoid repeated freeze-thaw cycles and store at -80°C. 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)
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Additional information
application information
Recommended dilution

1: 5000 (WB) 

Expected | apparent MW

12.5 | 14 kDa for A. thaliana

Confirmed reactivity

Arabidopsis thaliana

Predicted reactivity

cytc1 and cytc2 from following species: A. theoprasi, B. napus, B. oleracea, C. maxima, Ch. reinhardtii (peptide target partially conserved), L. esculentum, Lupinus luteus,  M. truncatula, Nicotiana tabacum, Oryza sativa, Ostreococcus (peptide target partially conserved), P. aurea, Physcomitrella patens, R. comunis, S. nigra, V. vinifera

Not reactive in

Arabidopsis thaliana CytC6

Additional information

The presence of cytochrome c in the cysotol is a marker of PCD (programmed cell death).

Selected references Schimmeyer et al. (2016). L-Galactono-1,4-lactone dehydrogenase is an assembly factor of the membrane arm of mitochondrial complex I in Arabidopsis. Plant Mol Biol. 2016 Jan;90(1-2):117-26. doi: 10.1007/s11103-015-0400-4. Epub 2015 Oct 31.
Li et al. (2016). Characterization of a novel β-barrel protein (AtOM47) from the mitochondrial outer membrane of Arabidopsis thaliana. J Exp Bot. 2016 Nov;67(21):6061-6075. Epub 2016 Oct 6.

Application example

western blot using affinity purified anti-cytochrome c antibodies
Mitochondrial proteins (15 ug) from Arabidopsis thaliana mitochondria was separated on 16% acrilamide gel and electrophoresis prepared according to Schägger and von Jagov (Anl. Biochem., 1987, 166:368-379). After running the gel, proteins were transferred to PVDF membrane using wet transfer (Roti®-Blot 2, Roth). Transfer was checked by Ponceau S staining. Blot was destained by several quick washings in distilled water and 1 washing in 1X TBS (10 mM T pH 7.5, 150 mM NaCl) (10-15 min.).Blot was blocked by 1.5 hour in 5% milk in TBST (1X TBS, 0,1% Tween 20) After blocking blot was washed quickly twice in TBST and incubated 2 hours with primary antibody (dilution 1: 1000) in TBST. Washing: two quick washings in TBST and 3 x 10 min. washings in TBST. Then blot was incubated 45-60 min. with a secondary anti-rabbit antibodies conjugated to peroxidase (Agrisera AB, dilution 1:10 000, AS09 602) in TBST. Washing: as above. After washing blot was incubated 1-2 min. in ECL solution (NOWA Kit, Thermo Scientific). Chemiluminescence was detected by BioSpectrum® Imaging System (UVP). Exposure time was 5 seconds.

Courtesy Dr. Janusz Piechota, Wrocław University, Poland


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