Development of a quantitative polyacrylamide gel electrophoresis analysis using a multichannel radioactivity counter for the evaluation of oligonucleotide-bound drug carrier.
A quantitative polyacrylamide gel electrophoresis (PAGE) analysis using a multichannel radioactivity counter was designed for the evaluation of 33P-labeled antisense oligonucleotide associated with polymeric drug carrier (nanoparticles). The proposed analytical method was first validated. The criteria of specificity, linearity, reliability, detection and quantification limits, and resolution power were determined. Results were compared to those obtained using liquid scintillation counting of crude samples or after solubilization of gel slices. The proposed method gave a better linearity and reliability than liquid scintillation counting of solubilized gel slices. In comparison with the liquid scintillation counting of crude samples, the method presented the advantage of being able to directly separate oligonucleotides differing by only one nucleotide in length. This method was applied for the separation of free oligonucleotides and oligonucleotides bound onto nanoparticles, allowing quantification of the amount of free and bound oligonucleotides without any further separation steps. Thus, because it is easy and rapid, the quantitative PAGE analysis using a multichannel radioactivity counter offers interesting possibilities for the characterization of oligonucleotide nanoparticles.
Aynié
I
I
Laboratoire de Physico-Chimie, Université Paris-Sud, Châtenay-Malabry, France.
Vauthier
C
C
Foulquier
M
M
Malvy
C
C
Fattal
E
E
Couvreur
P
P
D003487:Cyanoacrylates / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D004659:Enbucrilate; D007553:Isotope Labeling; D016376:Oligonucleotides, Antisense / Q000032:analysis* / Q000302:isolation & purification; D010761:Phosphorus Radioisotopes; D011108:Polymers / Q000737:chemistry; D015203:Reproducibility of Results; D012588:Scintillation Counting
D003487:Cyanoacrylates; D016376:Oligonucleotides, Antisense; D010761:Phosphorus Radioisotopes; D011108:Polymers; D004659:Enbucrilate