Demonstration of expression of six proteins of the mammalian cell entry (mce1) operon of Mycobacterium tuberculosis by anti-peptide antibodies, enzyme-linked immunosorbent assay and reverse transcription-polymerase chain reaction.
Polyclonal rabbit antibodies were generated to synthetic peptides corresponding to predicted B-cell epitopes of six proteins of the mce1 operon of Mycobacterium tuberculosis. Antipeptide antibodies reacted with Mce1A and Mce1E fusion proteins in sonicates of recombinant Escherichia coli as well as with distinct bands in sonicates, but not in culture fluids of M. tuberculosis and M. bovis bacillus Calmette-Guérin (BCG). Polyvalent rabbit antibodies generated by immunization with sonicates of BCG bacilli reacted with synthetic peptides from the six Mce1 proteins on the solid phase in enzyme-linked immunosorbent assay (ELISA), albeit with different frequencies. The Mce1A peptide (p124-140) reacted most frequently, with seven of the nine antibodies tested, while the Mce1F peptide (p329-343) reacted with two. Used as a control, 20 polyclonal rabbit antibodies to 12 isolated proteins of M. tuberculosis and M. bovis BCG did not react with any of the six synthetic peptides, except in one case. mRNA expression of the six mce1A-mce1F genes of M. tuberculosis was demonstrated by reverse transcription-polymerase chain reaction (RT-PCR). These data indicate that all Mce1A-Mce1F proteins of the mce1 operon are expressed by in vitro-grown M. tuberculosis and M. bovis BCG.
Harboe
M
M
Institute of Immunology, University of Oslo, The National Hospital, N-0027 Oslo, Norway.
Christensen
A
A
Haile
Y
Y
Ulvund
G
G
Ahmad
S
S
Mustafa
A S
AS
Wiker
H G
HG
D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial / Q000235:genetics; D001402:B-Lymphocytes / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics* / Q000276:immunology*; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009163:Mycobacterium bovis / Q000235:genetics / Q000276:immunology; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000276:immunology*; D009876:Operon; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D017931:DNA Primers; D000939:Epitopes; D011993:Recombinant Fusion Proteins