Counting low-copy number proteins in a single cell.
We have designed a microfluidic device in which we can manipulate, lyse, label, separate, and quantify the protein contents of a single cell using single-molecule fluorescence counting. Generic labeling of proteins is achieved through fluorescent-antibody binding. The use of cylindrical optics enables high-efficiency (approximately 60%) counting of molecules in micrometer-sized channels. We used this microfluidic device to quantify beta2 adrenergic receptors expressed in insect cells (SF9). We also analyzed phycobiliprotein contents in individual cyanobacterial cells (Synechococcus sp. PCC 7942) and observed marked differences in the levels of specific complexes in cell populations that were grown under nitrogen-depleted conditions.
Huang
Bo
B
Department of Chemistry, Stanford University, Stanford, CA 94305-5080, USA.
Wu
Hongkai
H
Bhaya
Devaki
D
Grossman
Arthur
A
Granier
Sebastien
S
Kobilka
Brian K
BK
Zare
Richard N
RN
D000818:Animals; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins / Q000032:analysis*; D001433:Bacteriolysis; D002232:Carbocyanines; D002460:Cell Line; D003470:Culture Media; D005453:Fluorescence; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes; D006801:Humans; D007834:Lasers; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation; D044085:Microfluidics; D009584:Nitrogen / Q000378:metabolism; D055095:Optics and Photonics; D045524:Phycobilisomes / Q000378:metabolism; D010798:Phycocyanin / Q000032:analysis*; D018343:Receptors, Adrenergic, beta-2 / Q000032:analysis*; D046940:Synechococcus / Q000737:chemistry* / Q000254:growth & development / Q000378:metabolism
D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D002232:Carbocyanines; D003470:Culture Media; D005456:Fluorescent Dyes; D045524:Phycobilisomes; D018343:Receptors, Adrenergic, beta-2; C002497:allophycocyanin; C085321:cyanine dye 5; D010798:Phycocyanin; D009584:Nitrogen