AS08 368 Anti-APX | L-ascorbate peroxidase

Confirmed reactivity:	Arabidopsis thaliana, Armeria maritima, Brassica napus, Capsicum annuum, Citrus sp., Digitaria sanguinalis, Dionaea muscipula, Echinochla crus-galli, Iris pumila, Lathyrus sativus, Liquidambar formosana, Lupin sp., Manihot esculenta, Medicago sativa, Nicotiana tabacum thylakoid-bound APX, stromal APX; Oryza sativa, Panicum milaceum, Plumbago zeylanica, Schima superba, Silene vulgaris, Solanum lycopersicum, Spinacia oleracea (stromal APX, thylakoid-bound), Triticum aestivum
Predicted reactivity:	Brassica rapa subsp. oleifera Stromal APX; Glycine max, Glycine soja L-ascorbate peroxidase T, chloroplastic; Medicago truncatula thylakoid-bound APX; Mesembryanthemum crystallinum, Pisum sativum Chloroplast stromal ascorbate peroxidase 12; Solanum lycopersicum thylakoid-bound APX; Spinacia oleracea stromal APX; Theobroma cacao L-APX T isoform 3; Vitis vinifera Species of your interest not listed? Contact us
Not reactive in:	Algae, Helianthus annus, Marchantia polymorpha

5 to 20 μg of total leaf protein from Arabidopsis thaliana (left panel) and chloroplast fractions (thylakoids and soluble, right panel) was separated on 15% polyacrylamide gel with 6M urea and blotted on PVDF. Filters were blocked 1h with 5% BSA, incubated with anti-APX antibody (1: 2000, 1h) followed by incubation with secondary HRP-coupled anti rabbit antibody (1: 10 000, 1h). Signal was detected with chemiluminescence detection reagent. AS08 368 is reactive to thylakoid (tAPX, 38 kDa), stromal (sAPX, 33 kDa), peroxisomal (pAPX, 31 kDa) and cytoplasmic (cAPX1 + cAPX2, 25 kDa) forms of ascorbate peroxidases.

 

Total proteins of Arabidopsis thaliana leaves were extracted with 10 % TCA and precipitated. The pellet was washed with acetone and resuspended in 100mM Tris-HCl (pH 7.5), 1mM EDTA, 2% (w= v) SDS, 1:100 of protease inhibitor cocktail (Thermo Scientific), 1 mM PMSF. Leaves were also grinded in 100 mM Tris-HCl (pH 7.5), MgCl2 10 mM, 1 mM EDTA, 1 mM PMSF, 1/100 of protease inhibitor cocktail and centrifugated. The supernatant (soluble fraction) was separated and the pellet (membrane fraction) was resuspended in the same buffer with 6 M urea and 1% SDS. Different amounts of proteins were separated in 15 % polyacrylamide gel with 6M urea after denaturation (70°C 5 min) and blotted on PVDF. Filters were blocked 1h with 5% BSA, Incubated with anti-APX antibodies at a dilution 1:2000, 1h/RT, washed 4 times with TBS tween (5 min each) and incubated with HRP coupled anti-rabbit IgG secondary antibody in dilution 1:10 000 1h/RT (AS09 602, Agrisera). After incubation with secondary antibody, the filter was washed 4 times with TBS (5 min each) and signal was detected with chemiluminescent detection reagent (30 secs exposition in film).

Courtesy Manuel Guinea Diaz, University of Turku, Finland

 

Reactant: Manihot esculenta (Cassava)

Application: Western Blotting

Pudmed ID: 24727655

Journal: PLoS One

Figure Number: 5B

Published Date: 2014-04-15

First Author: An, F., Fan, J., et al.

Impact Factor: 2.942

Open Publication

Western blotting of Rubisco, APX and PrxQ.The expression of Rubisco, APX and PrxQ in leaves of cassava NZ199 diploid (a) and autotetraploid (b) genotypes were detected by western blotting using antiRubisco-polyclonal antibody (AS07218), anti-APX antibody (AS08368) and anti-PrxQ antibody (AS05093) from Agrisera, respectively.

 

Reactant: Liquidambar formosana (Chinese sweet gum)

Application: Western Blotting

Pudmed ID: 25025692

Journal: PLoS One

Figure Number: 6A

Published Date: 2014-07-16

First Author: Chen, J., Hu, W. J., et al.

Impact Factor: 2.942

Open Publication

Western blot analysis showing the expression of three protein spots.(A) Expression of rubulose-1,5-bisphoshate carboxylase large subunit (RuBisco LSU), ascorbate peroxidase (APX) and glutathione-S-transferase (GST) in L. formosana and S. superba seedlings after AR treatment. Relative expression level of RuBisco LSU (B), APX (C) and GST (D) were analyzed with the Quantity One software. ?-actin was used as the internal control. Means with different letters indicate significantly difference (p<0.05) with regard to AR treatments.

 

Reactant: Arabidopsis thaliana (Thale cress)

Application: Western Blotting

Pudmed ID: 30609769

Journal: Int J Mol Sci

Figure Number: 4E

Published Date: 2019-01-02

First Author: Szyma?ska, K. P., Polkowska-Kowalczyk, L., et al.

Impact Factor: 5.542

Open Publication

SnRK2.4 and SnRK2.10 regulate enzymes of the ascorbate cycle during response to salt stress. Wild type and snrk2 mutant seedlings were subjected to treatment with 150 mM NaCl for times indicated. Expression of (A) APX1—Ascorbate Peroxidase 1, (B) APX2—Ascorbate Peroxidase 2, (C) APX6—Ascorbate Peroxidase 6, and (D) DHAR1—Dehydroascorbate Reductase 1 was monitored by RT-qPCR; error bars represent SD; stars represent statistically significant differences in comparison with the wt plants (Student t-test) where * p < 0.05; ** p < 0.001; *** p < 0.0001. Total protein level of (E) APX and (F) DHAR1 was monitored with immunoblot analysis; after exposure, membranes were stripped and reused for GAPDH detection as a loading control. At least two independent biological replicates of the experiment were performed. Results of one representative experiment are shown.

 

Reactant: Pisum sativum (Pea)

Application: Western Blotting

Pudmed ID: 33445673

Journal: Int J Mol Sci

Figure Number: 3A

Published Date: 2021-01-12

First Author: Tokarz, K. M., Weso?owski, W., et al.

Impact Factor: 5.542

Open Publication

Content of antioxidant enzymes: (a) APX isoforms and (b) CAT in leaves and stems of grass pea shoots under salinity; tAPX—thylakoid ascorbate peroxidase, sAPX—stromal ascorbate peroxidase, pAPX—peroxisomal ascorbate peroxidase, cAPX—cytoplasmic ascorbate peroxidise, CAT—catalase; content of proteins expressed as relative units [RU]; different letters—statistically significant differences within each organ (leaf-uppercase, stem-lowercase) at p ? 0.05 (n = 3).

This product can be sold containing proclin if requested

APX plays a key role in plant antioxidant system by reducing hydrogen peroxide to water. Cellular localization includes chloroplast (tAPX and sAPX), cytosol (cAPX) and peroxisome (pAPX).

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Agrisera Western Blot protocol and video tutorials

Protocols to work with plant and algal protein extracts


Oxygenic photosynthesis poster by prof. Govindjee and Dr. Shevela

Z-scheme of photosynthetic electron transport by prof. Govindjee and Dr. Björn and Dr. Shevela