Anti-L10 | Ribosomal protein L10 family protein

Samples:
1 – 30 µg of Arabidopsis thaliana WT mitochondria (10-week-old-plants, SD)
2 - 30 µg of Arabidopsis thaliana rps10 mitochondria (10-week-old-plants, SD)
3 – 15 µg of Arabidopsis thaliana WT mitochondria (10-week-old-plants, SD)
4 - 15 µg of Arabidopsis thaliana rps10 mitochondria (10-week-old-plants, SD)
5- 30 µg of Arabidopsis thaliana WT mitochondria (10-week-old-plants, SD)
6 - 30 µg of Arabidopsis thaliana rps10 mitochondria (10-week-old-plants, SD)
7--15 µg of Arabidopsis thaliana WT mitochondria (10-week-old-plants, SD)
8 - 15 µg of Arabidopsis thaliana rps10 mitochondria (10-week-old-plants, SD)
9- Marker
10 –30 µg of Arabidopsis thaliana WT leaf extract (4-day-old seedlings, constant light)

Mitochondria/total protein extract isolated freshly from Arabidopsis thaliana rossette leaves/seedlings. Samples were separated in the RT on 12 % SDS-PAGE and blotted for 2 h to PVDF in the RT. Blot was blocked with 10 % milk for 1h/RT with agitation. I blot was incubated in the primary antibody at a dilution of 1: 3000 for 1h/RT in PBS-T with agitation. II blot was incubated in the primary antibody at a dilution of 1: 3000 for O/N in TBS-T with agitation. The antibody solution was decanted and the blot was rinsed briefly, then washed once for 15 min and 3 times for 5 min in PBS-T/TBS-T at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated) diluted to 1: 25000 in PBS-T or TBS-T with 5 % milk for 1 h/RT with agitation. The blot was washed as above and developed with a following chemiluminescent detection reagent. Exposure time was 2 minutes.
Note:
Using more concetrated primary antibody combined with higher protein load/well and conditions which increased L10 levels, may contribute to detection of L10 in endogenous samples.

Courtesy of Dr. Małgorzata Kwaśniak-Owczarek, University of Wrocław, Polan