A novel 111In-labeled antisense DNA probe with multi-chelating sites (MCS-probe) showing high specific radioactivity and labeling efficiency.
A multi-aminolinked oligodeoxynucleotide (ODN) was synthesized by substitution of dT with aminolinked dU in the sequence, following conjugation with isothiocyanobenzyl-EDTA (IBE) for 111In labeling. As a model target gene, the c-erbB-2 protooncogene was used. The probability of the number of aminolinked dU in the 20mer ODN was 5, but there were actually 3 and 4 in the selected antisense and sense ODNs, respectively. The IBE/ODN conjugation levels of probes with multi-chelating sites (MCS-probe) were 1.6 (antisense) and 2.4 (sense), more than 50 times higher than those of our previous studies using 5'-end aminolinked ODNs (IBE/ODN = 0.03). Labeling studies using the MCS-probe and 111In indicated that specific radioactivity as high as 48 MBq/nmol could be obtained with a labeling efficiency of over 90%. The 111In-antisense-MCS-probe could bound to sense ODN under physiological conditions, but the 111In-sense-MCA-probe could not. Thus, side-chain modification of ODN for metal labeling is considered to be useful for antisense techniques.
Fujibayashi
Y
Y
Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo, Japan. yfuji@pharm.kyoto-u.ac.jp
Yoshimi
E
E
Waki
A
A
Sakahara
H
H
Saga
T
T
Konishi
J
J
Yonekura
Y
Y
Yokoyama
A
A
D016375:Antisense Elements (Genetics); D016373:DNA, Antisense / Q000138:chemical synthesis* / Q000378:metabolism; D004492:Edetic Acid / Q000031:analogs & derivatives; D007205:Indium Radioisotopes; D017879:Isothiocyanates; D007553:Isotope Labeling; D015513:Oncogene Proteins; D012333:RNA, Messenger
D016375:Antisense Elements (Genetics); D016373:DNA, Antisense; D007205:Indium Radioisotopes; D017879:Isothiocyanates; D015513:Oncogene Proteins; D012333:RNA, Messenger; C119668:isothiocyanobenzyl ethylenediamine tetracetic acid; D004492:Edetic Acid