The following antibodies were used in IHC and IF: guinea pig anti-insulin (DAKO, #A0564, 1:1,000), rabbit anti-glucagon (Phoenix, #H-028–05, 1:2,000), rabbit anti Ki67 (Abcam, #ab16667, 1:200), rabbit anti-cleaved caspase 3 (Cell Signaling, #9661, 1:200), rat anti-BrdU (Abcam, #ab6326, 1:200), mouse anti-Nkx6–1 (Hybridoma Bank, #F55A12, 1:100) rabbit anti-Mafa (Bethyl, #IHC-00352, 1:100), rabbit anti-Nkx2–2 (Sigma, #HPA003468, 1:100), rabbit anti-ALDH1A3 (Novus, #NBP2–15339, 1:100) and rabbit anti-gastrin (Millipore, #256A, 1:200). The following antibodies were used for western blot: rabbit anti-cleaved caspase 3 (Cell Signaling, #9661, 1:1,000), rabbit anti-adipsin (Santa Cruz, #sc-50419, 1:1,000), chicken anti-C3a (Abcam, #ab48581, 1:1,000), rabbit anti-Dusp26 (Invitrogen, #PA5–22013, 1:1,000), rabbit anti-actin (Cell Signaling, #8456, 1:1,000) and mouse anti-FLAG (Sigma, #A8592, 1:1,000). Recombinant C3a (R&D) was used at a concentration of 100 nM. NSC-87877 compound was dissolved in PBS and used at 20 μM concentration.
Recombinant c3a
Manufactured by R&D Systems
Recombinant C3a is a laboratory product produced by R&D Systems. It is a recombinant form of the complement system protein C3a. C3a is a small polypeptide that is released as part of the complement cascade activation.
Automatically generated - may contain errors
Lab products found in correlation
Anti insulin, by Agilent Technologies (1 mentions)
Rabbit anti actin, by Cell Signaling Technology (1 mentions)
Rabbit anti mafa, by Fortis Life Sciences (1 mentions)
Rat anti brdu, by Abcam (1 mentions)
Rabbit anti cleaved caspase 3, by Cell Signaling Technology (1 mentions)
Rabbit anti ki67, by Abcam (1 mentions)
Mouse anti flag, by Merck Group (1 mentions)
2 protocols using recombinant c3a
1
Comprehensive Immunohistochemistry and Western Blot Antibody Panel
2
Osteogenic Differentiation of BMSCs
Check if the same lab product or an alternative is used in the 5 most similar protocols
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BMSCs were plated in culture media (α-MEM media + 10 % Hyclone FBS + 1 % PSG) at 2.0 × 104 cells/cm2 in 12-well plates. Upon reaching confluency, cultures were stimulated with osteogenic media (α-MEM media + 10 % Hyclone FBS + 1 % PSG + 50μg/mL ascorbic acid) for 5 days, media changed every other day. Cells were then cultured overnight in serum-deprived osteogenic media (α-MEM media +0.3 % Hyclone FBS + 1 % PSG + 50μg/mL ascorbic acid). On Day-6, cultures were stimulated for 2 h with serum-deprived osteogenic media supplemented with either vehicle-control or recombinant C3a (5, 10, or 20 ng/mL, R&D Systems). After the 2-h stimulation, supernatants were aspirated, and cells were washed to remove recombinant proteins. Fresh serum-deprived osteogenic media was added back to cultures for 5 h. Cell culture supernatants were then collected for ELISA protein assays, and cells were isolated for qRT-PCR gene expression analysis to assess pro-osteoclastic genes. In vitro osteoblast stimulation assay was performed in triplicate (technical replicate) cultures. Data are representative of two separate experiments.
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