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Actively targeted in vivo multiplex detection of intrinsic cancer biomarkers using biocompatible SERS nanotags.

Dinish US, Balasundaram G, Chang YT, Olivo M - Sci Rep (2014)

Bottom Line: However, nanotags without antibodies showed no detectable signal after 6 hours.This difference could be due to the specific binding of the bioconjugated nanotags to the receptors on the cell surface.Thus, this study establishes SERS nanotags as an ultrasensitive nanoprobe for the multiplex detection of biomarkers and opens up its potential application in monitoring tumor progression and therapy and development into a theranostic probe.

View Article: PubMed Central - PubMed

Affiliation: 1] Singapore Bioimaging Consortium, Agency for Science Technology and Research (A*STAR), 11 Biopolis Way, Singapore 138667 [2].

ABSTRACT
Surface-enhanced Raman scattering (SERS) technique is becoming highly popular for multiplex biosensing due to the 'fingerprint' Raman spectra from every molecule. As a proof-of-concept, we demonstrated the actively targeted multiplex in vitro and in vivo detection of three intrinsic cancer biomarkers - EGFR, CD44 and TGFβRII in a breast cancer model using three multiplexing capable, biocompatible SERS nanoparticles/nanotags. Intra-tumorally injected antibody conjugated nanotags specifically targeting the three biomarkers exhibited maximum signal at 6 hours and no detectable signal at 72 hours. However, nanotags without antibodies showed no detectable signal after 6 hours. This difference could be due to the specific binding of the bioconjugated nanotags to the receptors on the cell surface. Thus, this study establishes SERS nanotags as an ultrasensitive nanoprobe for the multiplex detection of biomarkers and opens up its potential application in monitoring tumor progression and therapy and development into a theranostic probe.

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Related in: MedlinePlus

(A) Confirmation of the expression of intrinsic biomarkers-EGFR, CD44 and TGFβRII in 10 (a) and 15 μg (b) of total protein extracted from cultured MDA-MB-231 by western blot. Total proteins from MDA-MB-231 cells were electrophoresed under same experimental conditions and probed for β-actin, CD44 and TGFβRII proteins using respective primary antibodies followed by corresponding secondary antibodies. EGFR was detected by electrophoresing total proteins in SDS free conditions followed by probing using corresponding primary and secondary antibodies. (B) Cell viability study using Cell Counting Kit-8 on MDA-MB-231 cells exposed to pure AuNP, antibody-bioconjugated (w ab) and non bioconjugated (w/o ab) SERS nanotags (MGITC, Cy5 and Rh6G) after 24 hours.
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f2: (A) Confirmation of the expression of intrinsic biomarkers-EGFR, CD44 and TGFβRII in 10 (a) and 15 μg (b) of total protein extracted from cultured MDA-MB-231 by western blot. Total proteins from MDA-MB-231 cells were electrophoresed under same experimental conditions and probed for β-actin, CD44 and TGFβRII proteins using respective primary antibodies followed by corresponding secondary antibodies. EGFR was detected by electrophoresing total proteins in SDS free conditions followed by probing using corresponding primary and secondary antibodies. (B) Cell viability study using Cell Counting Kit-8 on MDA-MB-231 cells exposed to pure AuNP, antibody-bioconjugated (w ab) and non bioconjugated (w/o ab) SERS nanotags (MGITC, Cy5 and Rh6G) after 24 hours.

Mentions: MDA-MB-231 breast cancer cell line has been known to express a variety of cell surface receptors including the three cancer biomarkers -EGFR, CD44 and TGFβRII. To determine the expression levels of these proteins, western blot was performed on two different concentrations of total protein extracted from cultured MDA-MB-231 cells. All three proteins showed detectable levels of expression with EGFR (132 kDa) being highly expressed, CD44 (82 kDa) being second highly expressed while TGFβRII (68 kDa) being lowly expressed compared to the other two (Fig. 2A). The full blots of the cropped images represented here are given in Fig. S4 in SI. For targeted SERS detection of these biomarkers, each of the nanotags was appropriately conjugated to the antibodies against these proteins. For instance, EGFR with highest expression was linked to Rh6G nanotag having least SERS signal while CD44 and TGFβRII with lesser expression levels were conjugated respectively to MGITC and Cy5 with relatively higher SERS signal.


Actively targeted in vivo multiplex detection of intrinsic cancer biomarkers using biocompatible SERS nanotags.

Dinish US, Balasundaram G, Chang YT, Olivo M - Sci Rep (2014)

(A) Confirmation of the expression of intrinsic biomarkers-EGFR, CD44 and TGFβRII in 10 (a) and 15 μg (b) of total protein extracted from cultured MDA-MB-231 by western blot. Total proteins from MDA-MB-231 cells were electrophoresed under same experimental conditions and probed for β-actin, CD44 and TGFβRII proteins using respective primary antibodies followed by corresponding secondary antibodies. EGFR was detected by electrophoresing total proteins in SDS free conditions followed by probing using corresponding primary and secondary antibodies. (B) Cell viability study using Cell Counting Kit-8 on MDA-MB-231 cells exposed to pure AuNP, antibody-bioconjugated (w ab) and non bioconjugated (w/o ab) SERS nanotags (MGITC, Cy5 and Rh6G) after 24 hours.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3921631&req=5

f2: (A) Confirmation of the expression of intrinsic biomarkers-EGFR, CD44 and TGFβRII in 10 (a) and 15 μg (b) of total protein extracted from cultured MDA-MB-231 by western blot. Total proteins from MDA-MB-231 cells were electrophoresed under same experimental conditions and probed for β-actin, CD44 and TGFβRII proteins using respective primary antibodies followed by corresponding secondary antibodies. EGFR was detected by electrophoresing total proteins in SDS free conditions followed by probing using corresponding primary and secondary antibodies. (B) Cell viability study using Cell Counting Kit-8 on MDA-MB-231 cells exposed to pure AuNP, antibody-bioconjugated (w ab) and non bioconjugated (w/o ab) SERS nanotags (MGITC, Cy5 and Rh6G) after 24 hours.
Mentions: MDA-MB-231 breast cancer cell line has been known to express a variety of cell surface receptors including the three cancer biomarkers -EGFR, CD44 and TGFβRII. To determine the expression levels of these proteins, western blot was performed on two different concentrations of total protein extracted from cultured MDA-MB-231 cells. All three proteins showed detectable levels of expression with EGFR (132 kDa) being highly expressed, CD44 (82 kDa) being second highly expressed while TGFβRII (68 kDa) being lowly expressed compared to the other two (Fig. 2A). The full blots of the cropped images represented here are given in Fig. S4 in SI. For targeted SERS detection of these biomarkers, each of the nanotags was appropriately conjugated to the antibodies against these proteins. For instance, EGFR with highest expression was linked to Rh6G nanotag having least SERS signal while CD44 and TGFβRII with lesser expression levels were conjugated respectively to MGITC and Cy5 with relatively higher SERS signal.

Bottom Line: However, nanotags without antibodies showed no detectable signal after 6 hours.This difference could be due to the specific binding of the bioconjugated nanotags to the receptors on the cell surface.Thus, this study establishes SERS nanotags as an ultrasensitive nanoprobe for the multiplex detection of biomarkers and opens up its potential application in monitoring tumor progression and therapy and development into a theranostic probe.

View Article: PubMed Central - PubMed

Affiliation: 1] Singapore Bioimaging Consortium, Agency for Science Technology and Research (A*STAR), 11 Biopolis Way, Singapore 138667 [2].

ABSTRACT
Surface-enhanced Raman scattering (SERS) technique is becoming highly popular for multiplex biosensing due to the 'fingerprint' Raman spectra from every molecule. As a proof-of-concept, we demonstrated the actively targeted multiplex in vitro and in vivo detection of three intrinsic cancer biomarkers - EGFR, CD44 and TGFβRII in a breast cancer model using three multiplexing capable, biocompatible SERS nanoparticles/nanotags. Intra-tumorally injected antibody conjugated nanotags specifically targeting the three biomarkers exhibited maximum signal at 6 hours and no detectable signal at 72 hours. However, nanotags without antibodies showed no detectable signal after 6 hours. This difference could be due to the specific binding of the bioconjugated nanotags to the receptors on the cell surface. Thus, this study establishes SERS nanotags as an ultrasensitive nanoprobe for the multiplex detection of biomarkers and opens up its potential application in monitoring tumor progression and therapy and development into a theranostic probe.

Show MeSH
Related in: MedlinePlus