IBB PAS Repository

Effect of unlabeled helper probes on detection of an RNA target by bead-based sandwich hybridization

Barken, Kim Bundvig and Gabig-Cimińska, Magdalena and Holmgren, Anders and Molin, Søren (2004) Effect of unlabeled helper probes on detection of an RNA target by bead-based sandwich hybridization. Biotechniques, 36 (1). pp. 124-132. ISSN 0736-6205


Official URL: http://www.biotechniques.com/archive/


Unlabeled helper oligonucleotides assisting a bead-based sandwich hybridization assay were tested for the optimal placement of the capture and detection probes. The target used was a full-length in vitro synthesized mRNA molecule. Helper probes complementary to regions adjacent to the binding site of the 5' end attached capture probe were found much more effective than helper probes targeting positions adjacent to the detection probe binding site. The difference is believed to be caused by a disruption of the RNA secondary structure in the area where the capture probe binds, thereby reducing structural interference from the bead. The use of additional helpers showed an additive effect. Using helpers at both sides of the capture and detection probes showed a 15- to 40-fold increase in hybridization efficiency depending on the target, thereby increasing the sensitivity of the hybridization assays. Using an electrical chip linked to the detection probe for the detection of p-aminophenol, which is produced by alkaline phosphatase, a detection limit of 2 x 10(-13) M mRNA molecules was reached without the use of a nucleic acid amplification step.

Item Type:Article
Subjects:Q Science > QH Natural history > QH301 Biology
Q Science > QH Natural history > QH426 Genetics
Divisions:Laboratory of Molecular Biology (in Gdansk)
ID Code:1071
Deposited By: PhD Piotr Golec
Deposited On:17 Dec 2015 10:09
Last Modified:17 Dec 2015 10:09

Repository Staff Only: item control page