BD LSR II Cell Analyzer

BD LSR II Cell Analyzer

3-laser, 11 color (6-3-2)

BD LSR II Configuration 3-laser, 11 color
Excitation Laser	Detection Filter	example
488nm Blue Laser	530/30	Alexa Fluor 488, FITC, BB515
	575/26	PE
	610/20	PE-Texas Red
	660/20	PE-Cy5
	695/40	PI, PerCP, PerCP-Cy5.5, BB700, 7-AAD
	780/60	PE-Cy7
633nm Red Laser	670/30	APC, Alexa Fluor 647
	730/45	Alexa Fluor 700
	780/60	APC-Cy7, APC-H7
405nm Violet Laser	450/50	Alexa Fluor 405, V450, BV421, PB
405nm Violet Laser	525/50	Alexa Fluor 430, V500, BV510

Consumables

(1) 5-mL polystyrene test tubes, 12 x 75-mm (BD Falcon Cat. No.352052)

1. Sample preparation

(1) Recommend sample concentration: 1x10⁶cells /ml

(2) Filter your samples

Recommend filtering samples through a nylon mesh 40 µm (BD Cat. No. 352340) or 5-mL polystyrene test tubes, 12 x 75-mm (BD Falcon) with cell-strainer cap (BD Cat. No. 352235) if aggregated cells can be seen in the sample tube.

Filtered sample should be saved on ice, protected from light.

Data

8-color panel for Antigenspecific Phenotyping

Using an eight-color panel for antigenspecific phenotyping, informational content is maximized providing possible evidence of novel patterns of T-cell differentiation among CMV-responsive donors. Figure B1 shows a sequential gating strategy for phenotyping of IFN_ positive cells after CMV simulations. Figure B2 are the results of total CD8 T cells from a CMV+ donor.

Immunophenotyping

Characterizing cells at different stages of development through the use of fluorescent-labeled monoclonal antibodies against surface markers is one of the most common applications of flow cytometry. Surface staining and intracellular staining are common application for flow cytometry. PBMCs stimulated with SEB and protein transport inhibitor -BFA and detect the expression of IFN-γ.

Attachment:

1. BD LSRII User Guide

2. FACSDivav6_LSRII_quickguide

Reference website: BD Bioscience

https://www.bd.com/resource.aspx?IDX=17868

http://www.indiana.edu/~fccf/pdf/LSRIIBrochure.pdf