ABCMdb

PMID: 11839251

Connolly L, Moran E, Larkin A, Scheffer G, Scheper R, Sarkadi B, Kool M, Clynes M
A new monoclonal antibody, P2A8(6), that specifically recognizes a novel epitope on the multidrug resistance-associated protein 1 (MRP1), but not on MRP2 nor MRP3.
Hybrid Hybridomics. 2001;20(5-6):333-41., [PubMed]

Sentences

No. Mutations Sentence Comment

43 ABCC2 p.Leu23Ser ABCC2 p.Leu23Arg Control cell lines The multidrug resistant nonsmall cell lung carcinoma cell line, COR-L23R (made resistant by exposure to increasing concentrations of adriamycin(26)) and its drug-sensitive counterpart, COR-L23S, were used as control cells for the presence/absence of MRP1. Login to comment 44 ABCC2 p.Leu23Ser ABCC2 p.Leu23Arg Overexpression of MRP1 in COR-L23R cells compared with the parental COR-L23S had been confirmed in previous studies using other commercially available antibodies raised against MRP1 (MRPr1, MRPm5, MRPm6,(15) QCRL-1, and QCRL-3(16)) (Alexis Biochemicals, San Diego, CA). Login to comment 56 ABCC2 p.Leu23Arg These supernatants were also screened by Western blot analysis, CONNOLLY ET AL.334 against cell membrane preparations of the MRP1-overexpressing COR-L23R cell line. Login to comment 76 ABCC2 p.Leu23Arg MAb P2A8(6) detects a 190-kDa band (associated with the presence of MRP1) on preparations from drug resistant cells (COR-L23R) and MRP1-transfected cells (2008 MRP1). Login to comment 77 ABCC2 p.Leu23Ser P2A8(6) also detects a very faint 190-kDa band on preparationsfrom drug-sensitivecells (COR-L23S) and non- transfectedcells (2008 parental)indicatingan intrinsic basal expression of MRP1. Login to comment 88 ABCC2 p.Leu23Arg The antibody-peptide complex was centrifuged at 15,000 3 g in a microcentrifuge (Heraeus Instruments, Hanau, Germany) and the supernatantremoved carefully before use in Western blotting studies on COR-L23R cells. Login to comment 91 ABCC2 p.Leu23Arg Heat treatment appears to alter the expression of the antigen recognizedby MAb MRPr1 on the MRP1 protein expressed by the drug-resistant cells (COR-L23R). Login to comment 96 ABCC2 p.Leu23Arg Heat treatment does not appear to destroy the antigen recognized by MAb PA8(6) on the MRP1-drug resistant cells (COR-L23R) or MRP1-transfected cells (2008 MRP1). Login to comment 102 ABCC2 p.Leu23Arg MAb P2A8(6), like the well-characterized MRPr1 MAb, detects a 190-kDa band (associatedwith the presence of MRP1) in the COR-L23R and MRP1-transfected 2008 MRP1 cell lines. Login to comment 103 ABCC2 p.Leu23Arg These antibodies also detect a very low basal level of MRP1 expressed in the parental sensitiveCOR-L23R and parental nontransfected2008 cell lines. Login to comment 109 ABCC2 p.Leu23Arg Both MAbs detect MRP1 in the COR-L23R, HL60ADR, A549 and RPMI 2650 melphalan-selected cells. Login to comment 110 ABCC2 p.Leu23Ser A very low basal level of MRP1 is detected in the sensitive COR-L23S cells. Login to comment 115 ABCC2 p.Leu23Ser ABCC2 p.Leu23Arg MRP2 is undetectable in the COR-L23R, COR-L23S, HL60ADR, HL60S, DLKP, RPMI 2650, and RPMI 2650 taxol-selected cells. Login to comment 117 ABCC2 p.Leu23Arg MRP3 Heat treatment studies were carried out on the MRP1 overexpressing drug selected COR-L23R cell line and the MRP1-transfected cell line 2008 MRP1 prior to electrophoresis (cell lysates in loading buffer, placed in a water-bath for 3 min @ 100°C) (Figs. Login to comment 119 ABCC2 p.Leu23Arg These studies showed that heat treatment appears to decrease antigen recognition (observed mainly by a decrease in stainingintensityin the lower area of the MRP1 protein band on the Western blot) by the MRPr1 antibody in the drug-selectedCOR-L23R cell line. Login to comment 121 ABCC2 p.Leu23Arg The antigen recognized by antibody P2A8(6) appears to be heat stable in the drug selected COR-L23R cell line and the MRP1-transfected 2008 MRP1 cell line, as no decrease in staining intensity in the MRP1 protein band was observed following heat treatment (Fig. 4). Login to comment 124 ABCC2 p.Leu23Arg MRP1 expressionwas detectedby both these MAbs in the COR-L23R, HL60ADR, A549 and RPMI 2650 melphalan-selected cell lines. Login to comment 126 ABCC2 p.Leu23Ser Neither MAb detected MRP1 expression in the COR-L23S, HL60S, DLKP, DLKPA, or RPMI 2650 taxol-selected cell lines. Login to comment 134 ABCC2 p.Leu23Ser ABCC2 p.Leu23Arg SUMMARY OF MRP1, MRP2, AND MRP3 PROTEIN EXPRESSION LEVELS AS DETERMINED BY WESTERN BLOT ANALYSIS Protein expression levels (as determined by Western blot analysis) MRPr1 M2III-6 M3II-21 Cell line P2A8(6) (MRP1) (MRP2) (MRP3) COR-L23R 1 1 - - COR-L23S 1/2 1/2 - - HL60ADR 1 1 - - HL60S - - - - DLKP - - - - DLKPA - - - - A549 1 1 1 - RPMI 2650 - 1/2 - - Parental RPMI 2650 - - - - Taxol-selected RPMI 2650 1 1 1 - Melphalan-selected Key: -: Undetectable levels; 1: Over-expression; and 1/2: Very low basal levels. Login to comment 139 ABCC2 p.Leu23Arg Strong staining was seen on the non-Pgp MDR cell line COR-L23R and the MRP1-transfected 2008 ovarian carcinoma cell line but not their parental drug sensitive or nontransfected cell lines. Login to comment 141 ABCC2 p.Leu23Ser MAb P2A8(6) did not detect overexpressionof MRP1 in the sensitiveCOR-L23S, 2008 parental,2008 MRP2, or 2008 MRP3 cell lines but like MRPr1, P2A8(6) did detect a very low basal level of staining in these cell lines. Login to comment 146 ABCC2 p.Leu23Arg MAb P2A8(6) shows strong plasma-membranousstaining of the MRP1 overexpressingdrug selected cells (COR-L23R) and the MRP1-transfected cells (2008 MRP1). Login to comment