Raman nanoparticle probes for antibody-based protein detection in tissues.
Surface-enhanced Raman scattering (SERS) nanoparticles are emerging as a new approach for optical detection of biomolecules. In a model assay in formalin-fixed paraffin-embedded (FFPE) prostate tissue sections, we detect prostate-specific antigen (PSA) using antibody (Ab) conjugated to composite organic-inorganic nanoparticles (COINs), and we use identical staining protocols to compare COIN-Ab and Alexa-Ab conjugates in adjacent tissue sections. Spectral analysis illustrates the fundamental difference between fluorescence and Raman signatures and accurately extracts COIN probe signals from background autofluorescence. Probe signals are used to generate images of PSA expression on the tissue, and quality measures are presented to characterize the performance of the COIN assay in comparison to Alexa. Staining accuracy (ability to correctly identify PSA expression in epithelial cells) is somewhat less for COIN than Alexa, which is attributed to an elevated false negative rate of the COIN. However, COIN provided signal intensities comparable to Alexa, and good intra-, inter-, and lot-to-lot consistencies. Overall, COIN and Alexa detection reagents possess similar performance with FFPE tissues, supporting the further development of Raman probes for this application. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.
Lutz
Barry
B
Division of Public Health Sciences, Fred Hutchinson Cancer Research Center, M5-A864, 1212 Aloha St., Seattle, WA 95054, USA.
Dentinger
Claire
C
Sun
Lei
L
Nguyen
Lienchi
L
Zhang
Jingwu
J
Chmura
Aj
A
Allen
April
A
Chan
Selena
S
Knudsen
Beatrice
B
D000906:Antibodies; D005260:Female; D005404:Fixatives; D005456:Fluorescent Dyes; D005557:Formaldehyde; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D053758:Nanoparticles; D016612:Paraffin Embedding; D011467:Prostate / Q000378:metabolism; D017430:Prostate-Specific Antigen / Q000276:immunology / Q000378:metabolism; D011506:Proteins / Q000276:immunology / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013059:Spectrum Analysis, Raman / Q000379:methods*
D000906:Antibodies; D005404:Fixatives; D005456:Fluorescent Dyes; D011506:Proteins; D005557:Formaldehyde; D017430:Prostate-Specific Antigen