TY - GEN
T1 - QDot nanocrystals in intracellular flow
T2 - Nanotechnology 2010: Bio Sensors, Instruments, Medical, Environment and Energy - 2010 NSTI Nanotechnology Conference and Expo, NSTI-Nanotech 2010
AU - Sotnikov, Ilya
AU - Jaron, Shulamit
AU - Konopleva, Marina
AU - O'Brien, Susan
AU - Andreeff, Michael
AU - Hillabrant, Julia
AU - Manis, John
AU - Brown, Jennifer
AU - Vorobjev, Ivan
AU - Barteneva, Natasha
N1 - Copyright:
Copyright 2010 Elsevier B.V., All rights reserved.
PY - 2010
Y1 - 2010
N2 - We present for the first time a methodological approach for the measurement of cytoplasmic Zap-70 and nuclear Ki-67 proteins with QDot nanocrystal-labeled antibodies. Mononuclear cells from chronic lymphoid leukemia patients were used to detect Zap-70, and Jurkat T-cells for Ki-67. Intracellular staining kits based on saponin and paraformaldehyde were chosen as optimal for the delivery of QDot nanocrystals inside cells. Single staining with QDot 565-, 605-, or 655-labeled antibodies is operative in intracellular flow cytometry. In multicolor panels, the most robust results are obtained with the QDot 655 detected in the APC channel (633 nm excitation, 660/20 nm band pass or customized 655/20 filter) to simplify the spectral overlap compensation. In 3-, 4-, and 5-color combinations signal brightness for intracellular QDot 655 does not decrease, whereas positive-to-negative signal ratio is higher than with organic dyes. The optimized combination of fluorophores for 5-color panel includes APC-Cy7, PE, PE-Cy7, FITC, and QDot 655. We used CD3, CD19, CD38, and CD5 surface markers. Intracellular QDots outperformed organic fluorophores probed in the same samples. QDot nanocrystals are a robust tool in intracellular flow cytometry due to their narrow emission spectrum, brightness, and stability of this fluorophore.
AB - We present for the first time a methodological approach for the measurement of cytoplasmic Zap-70 and nuclear Ki-67 proteins with QDot nanocrystal-labeled antibodies. Mononuclear cells from chronic lymphoid leukemia patients were used to detect Zap-70, and Jurkat T-cells for Ki-67. Intracellular staining kits based on saponin and paraformaldehyde were chosen as optimal for the delivery of QDot nanocrystals inside cells. Single staining with QDot 565-, 605-, or 655-labeled antibodies is operative in intracellular flow cytometry. In multicolor panels, the most robust results are obtained with the QDot 655 detected in the APC channel (633 nm excitation, 660/20 nm band pass or customized 655/20 filter) to simplify the spectral overlap compensation. In 3-, 4-, and 5-color combinations signal brightness for intracellular QDot 655 does not decrease, whereas positive-to-negative signal ratio is higher than with organic dyes. The optimized combination of fluorophores for 5-color panel includes APC-Cy7, PE, PE-Cy7, FITC, and QDot 655. We used CD3, CD19, CD38, and CD5 surface markers. Intracellular QDots outperformed organic fluorophores probed in the same samples. QDot nanocrystals are a robust tool in intracellular flow cytometry due to their narrow emission spectrum, brightness, and stability of this fluorophore.
KW - Cytometry
KW - Ki-67
KW - Quantum dots
KW - ZAP-70
UR - http://www.scopus.com/inward/record.url?scp=78049451931&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=78049451931&partnerID=8YFLogxK
M3 - Conference contribution
AN - SCOPUS:78049451931
SN - 9781439834152
T3 - Nanotechnology 2010: Bio Sensors, Instruments, Medical, Environment and Energy - Technical Proceedings of the 2010 NSTI Nanotechnology Conference and Expo, NSTI-Nanotech 2010
SP - 424
EP - 427
BT - Nanotechnology 2010
Y2 - 21 June 2010 through 24 June 2010
ER -