NRF1 and NRF2 mRNA and Protein Expression Decrease Early during Melanoma Carcinogenesis: An Insight into Survival and MicroRNAs

Table 2

Antibodies used in immunohistochemical staining and western blot.

Immunohistochemistry
Antigen	1^st antibody	Dilution	Revealing of antigen	Incubation	Detection of 1^st AB	Colour development	Product	Clone
NRF1	Rabbit polyclonal anti-NRF1; (C-terminal)	1 : 500	Tris-EDTA; pH 9.0	2 h RT	Dako EnVision	DAB	Sigma, HPA065424	Not reported
NRF1	Mouse monoclonal anti-NRF1; (N-terminal)	1 : 500	10 mM sodium citrate buffer; pH 6.0	45 min RT	Dako EnVision	Fast Red	Santa Cruz Biotechnology, sc-365651	E-4
NRF2	Rabbit monoclonal anti-NRF2 phospho S40	1 : 600	10 mM sodium citrate buffer; pH 6.0	30 min RT	Dako EnVision	DAB	Abcam, 76086	EP1809Y
Western blot
Antigen	1^st antibody	Dilution					Product	Clone
NRF1	Rabbit polyclonal anti-NRF1; (C-terminal)	1 : 2500					Sigma, HPA065424	Not reported
NRF1	Mouse monoclonal anti-NRF1; (N-terminal)	1 : 1000					Santa Cruz Biotechnology, sc-365651	E-4
NRF2	Rabbit monoclonal anti-NRF2 phospho S40	1 : 10000					Abcam, 76086	EP1809Y
β-Actin	Mouse monoclonal anti-β-actin	1 : 5000					Novus Biologicals, NB600-501SS	AC-15
PCNA	Mouse monoclonal anti-PCNA	1 : 2000					Cell Signalling Technology, #2586	PC10
	HRP-conjugated goat-anti-mouse	1 : 5000					Santa Cruz Biotechnology, sc-2055	—
	HRP-conjugated goat-anti-rabbit	1 : 5000					Santa Cruz Biotechnology, sc-2054	—

RT = room temperature, DAB = diaminobenzidine.