Surface Enhanced Raman Scattering Based in Situ Hybridization Strategy for Telomere Length Assessment

Abstract

Assessing telomere length is of vital importance since telomere length is closely related with several fatal diseases such as atherosclerosis and cancer. Here, we present a strategy to assess/measure telomere length, that is, surface enhanced Raman scattering (SERS) based in situ hybridization (SISH). The SISH method uses two kinds of SERS nanoprobes to hybridize in situ with telomeres and centromeres, respectively. The telomere specific SERS nanoprobe is called the Telo-probe, while the centromere specific SERS nanoprobe is called the Centro-probe. They are composed of metal nanoparticles (NPs), Raman reporter molecules and specially designed DNA strands. With longer telomeres, more Telo-probes will hybridize with them, resulting in a stronger SERS signal. To exclude possible influence of the SERS intensity by external factors (such as the nanoprobe concentration, the cell number or different batches of nanoprobes), centromeres are used as the inner control, which can be recognized by Centro-probes. Telomere length is evaluated using a redefined telomere-to-centromere ratio (T/C ratio). The calculation method for T/C ratio in SISH method is more reliable than that in fluorescent in situ hybridization (FISH). In addition, unlike FISH method, the SISH method is insensitive to autofluorescence. Moreover, SISH method can be used to analyze single telomeres. These features make SISH an excellent alternative strategy for telomere length measurement.

Combining Multiplex SERS Nanovectors and Multivariate Analysis for In Situ Profiling of Circulating Tumor Cell Phenotype Using a Microfluidic Chip

“…SERS Aptamer Nanovector Synthesis : Sphere AuNPs were synthesized according to the standard sodium citrate‐reduced method . The concentration of AuNP was ≈3.3 × 10 −9 m .…”

“…SERS Aptamer Nanovector Synthesis: Sphere AuNPs were synthesized according to the standard sodium citrate-reduced method. [23] The concentration of AuNP was ≈3.3 × 10 −9 m. Thiol-modified DNA aptamers (with ten T bases elongated) were conjugated onto AuNP following a previously described method. [24] Briefly, 20 µL DNA (in 10 × 10 −3 m phosphate buffered saline (PBS)) was added to 1 mL of gold nanocolloid solution and shaken gently.…”

“…Next, probes were encapsulated with a silver shell, which plays a role in enhancing SERS signals and avoiding interference to Raman reporters from cells or other latent chemical entities in the channel. [18,23,25] 30 µL solution of 10 × 10 −3 m ascorbic acid and 15 µL of 10 × 10 −3 m silver nitrate solution were mixed into the suspension successively under gently stirring at 25 °C. The color of solution would change into bright orange after several hours' reaction.…”

Nanoscale Properties of Human Telomeres Measured with a Dual Purpose X-ray Fluorescence and Super Resolution Microscopy Gold Nanoparticle Probe

“…AZT is a putative telomerase inhibitor and has been shown to shorten telomere lengths in human cells. 19 Using qPCR we measured the lengths of telomeres in the treated and untreated cells ( Figure 5 a). The results show that HEK cells have on average 10.2 ± 0.12 kbps long telomeres, whereas the HEK-AZT treated cells have on average 9.2 ± 0.12 kbps long telomeres, which is significantly different in a t test at P = 0.005.…”

A resonance Rayleigh scattering sensor for sensitive differentiation of telomere DNA length and monitoring special motifs (G-quadruplex and i-motif) based on the Ag nanoclusters and NAND logic gate responding to chemical input signals

“…Thus, the evaluation of telomere length is important in understanding clinical significance of the telomere. To date, there are plenty of methods to measure telomere length, including polymerase chain reaction [ 6 ], hybridization protection assay [ 7 ], situ hybridization [ 8 ], flow cytometry [ 9 ], primed in situ [ 10 ] and single telomere length analysis [ 11 ]. Nevertheless, these methods require a large amount of starting material (0.5–5 μg DNA), and specialized, expensive equipment.…”