Ultrasensitive molecular beacon (MB) DNA biosensors, with micrometer to sub
micrometer sizes, have been developed for DNA/RNA analysis. The fluorescenc
e-based biosensors have been applied in DNA/RNA detection without the meed
for a dye-labeled target molecule or an intercalation reagent in the testin
g solution. Molecular beacons are hairpin-shaped oligonucleotides that repo
rt the presence of specific nucleic acids. We have designed a surface-immob
ilizable biotinylated ssDNA molecular beacon for DNA hybridization at a liq
uid-solid interface. The MBs have been immobilized onto ultrasmall optical
fiber probes through avidin-biotin binding. The MB DNA biosensor has been u
sed directly to detect, in real time, its target DNA molecules without the
need for a competitive assay. The biosensor is stable and reproducible. The
MB DNA biosensor has selectivity with single basepair mismatch identificat
ion capability. The concentration detection limits and mass detection limit
s are 0.3 nM and 15 amol for a 105-mu m biosensor, and 10 nM and 0.27 amol
for a submicrometer biosensor, respectively. We have also prepared molecula
r beacon DNA biosensor arrays for simultaneous analysis of multiple DNA seq
uences in the same solution. The newly developed DNA biosensors have been u
sed for the precise quantification of a specific rat gamma-actin mRNA seque
nce amplified by the polymerase chain reaction, (C) 2000 Academic Press.