Anti-Lhcb3 | LHCII type III chlorophyll a/b-binding protein

Cucumis melo, Dicots, Gymnosperms, Mosses

1 - 10 ug of 4 days old of wild-type Arabidopsis thaliana seedlings extract

2 - 5 ug of 4 days old of wild-type Arabidopsis thaliana seedlings extract

3- 10 ug of 4 days old of gun5-1 mutant seedlings extract

Courtesy of Dr .Duorong Xu, LMU München, Germany

From 1 μg to 8 μg of chlorophyll from Arabidopsis thaliana chloroplasts extracted with 0.4 M sorbitol, 50 mM Hepes NaOH, pH 7.8, 10 mM NaCl, 5 mM MgCl 2 and 2 mM EDTA were loaded to lanes. Samples were denatured with Laemmli buffer at 75 0 C for 5 min and were separated on 12% SDS-PAGE, and blotted 30 min to PVDF using wet transfer. Blot was blocked with 5% milk for 2h at room temperature (RT) with agitation. Blot was incubated in the primary antibody Anti-Lhcb3 (LOT 1901) at a dilution of 1: 2000 in 1% milk in TBS-T overnight at 4 0 C with agitation. The antibody solution was decanted and the blot was washed 4 times for 5 min in TBS-T at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG HRP conjugated, from Agrisera, AS09 602) diluted to 1:20 000 in 1 % milk in TBS-T for 1h at RT with agitation. The blot was washed 5 times for 5 min in TBS-T and 2 times for 5 min in TBS, and developed for 1 min with 1.25 mM luminol, 0.198 mM coumaric acid and 0.009% H ₂O₂ in 0.1 M Tris- HCl, pH 8.5. Exposure time in ChemiDoc System was 240 seconds.

Courtesy of Dr. Wioleta Wasilewska-Dębowska, University of Warsaw, Poland

Antibody format is a total IgG fraction, which means that it is a pool of polyclonal antibodies obtained by purification of serum on Protein G, not on a specific antigen column.

The major light-harvesting antenna complex II (LHCII) in photsynthetic eukaryotes is located in the thylakoid membrane of the chloroplast. It is a heterotrimeric complex formed by up to 3 different individual subtypes of chlorophyll a/b-binding proteins: Lhcb1, Lhcb2, and Lhcb3. While Lhcb1 and Lhcb2 are quite similar and regularily present in multiple gene-copies, the Lhcb3 protein differs in pigment-composition and molecular size and often is coded by only a single gene. Lhcb3 seems not to be present in the mobile LHCII trimers involved in state 1-state 2 transitions.
A molecular characterisation of the LHCII proteins can be found in Caffarri et al. (2004) A Look within LHCII:  Differential Analysis of the Lhcb1−3 Complexes Building the Major Trimeric Antenna Complex of Higher-Plant Photosynthesis. Biochemistry 43 (29): 9467–9476.