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1. FLOW CYTOMETRY FACILITY
Flow cytometry is a powerful technique for the analysis of multiple parameters of individual cells within heterogeneous populations. The flow cytometers performs this analysis by passing thousands of cells per second through a laser beam and capturing the light that emerges from each cell as it passes through. The data gathered can be analyzed statistically by flow cytometry softwares to report cellular characteristics such as:
• Cell size (FSC)
• Cell complexity (SSC)
• Fluorescence
2. EQUIPMENT
The Flow Cytometry Core Facility of the Andalusian Center for Developmental Biology is equipped with the following instruments:
1. BD FACSAria Cell Sorter

• Two lasers: 488 nm y 633 nm
• 9 parameters: FSC, SSC and 7 fluorescence colors
• Sample acquisition with temperature control
• Acquisition rates of up to 70 000 events per seconds
• Sorts of up to four populations simultaneously
• Cell sorting in tube or culture plate
• Nozzles: 70 and 100 micron
• Software: FACSDiva
The available optic configuration is detailed below:
Laser |
PMT |
LP Mirror |
BP Filter |
Fluorocromos |
Blue 488 nm |
A |
735 |
780/60 |
PE-Cy7 |
B |
655 |
695/40 |
PerCP-Cy5.5 ó PI |
675/20 |
PerCP ó PE-Cy5 |
C |
595 |
610/20 |
PE-Texas Red |
D |
556 |
576/26 |
PE ó PI |
585/42 |
PE/PI |
E |
502 |
530/30 |
FITC |
F |
|
488/10 |
SSC |
Red 633 nm |
A |
735 |
780/60 |
APC-Cy7 |
B |
685 |
712/21 |
APC-Cy5.5 |
C |
660/20 |
660/20 |
APC |
2. BD FACSCalibur

• Laser 488
• 5 parameters: FSC, SSC and 3 fluorescence colors
• Cell Sorting
• Software: Cell Quest Pro
The available optic configuration is detailed below:
Laser |
PMT |
LP Mirror |
Filter |
Fluorochromes |
BLUE 488 nm |
FL1 |
|
530/30 |
FITC |
FL2 |
560 |
585/42 |
PE |
FL3 |
640 |
650 LP |
PerCP |
|
|
488/10 |
FSC, SSC |
3. SONY MA900 Cell Sorter

• Four lasers on two axes: 488/561nm and 405/638 nm
• 14 parameters: FSC, BSC and 12 fluorescence colours
• Sample acquisition tube types: 0,5ml, 1.5ml, 5ml and 15ml
• Acquisition rates of up to 70 000 events per second
• Temperature control of sample and collection tubes: 5
oC and 37
oC
• Cell sorting in 2-way and 4-way tubes, epp or 6,12,24,48,96 & 384- well plates
• Index sorting software
• Nozzles: 70 and 100 micron
• Software: SONY-MA (online/offline)
The available optic configuration is detailed below:
Laser |
Optical filters |
LPF |
BP Filter |
Fluorochromes |
405nm |
FL6 |
|
450/50 |
Dapi, Bviolet 421 |
FL7/LP10 |
487.5LP |
525/50 |
BViolet510,Pacific Orange |
FL8/LP4 |
561LP |
585/30 |
BViolet570 |
FL9/LP3 |
600LP |
617/30 |
BViolet 605 |
FL10/LP7 |
639LP |
665/30 |
BViolet 650 |
FL11/LP6 |
685LP |
720/60 |
BViolet 711 |
FL12/LP5 |
752LP |
785/60 |
BViolet 785 |
488nm |
FL1 |
|
525/50 |
Fitc, AlexaFluor 488,EGFP,EYFP |
FL2/LP4 |
561LP |
585/30 |
PE |
FL3/LP3 |
600LP |
617/30 |
PE-Texas Red, PI, |
FL4/LP7 |
639LP |
695/50 |
7AAD, PE-cy5, PerCP |
FL5/LP1 |
752LP |
785/60 |
PE-CY7 |
BSC |
|
488/17B |
|
561nm |
FL2/LP4 |
561LP |
585/30 |
mOrange, DsREd-Monomer, tdTomato |
FL3/LP3 |
600LP |
617/30 |
mCherry, Alexa Fluor 594 |
FL4/LP7 |
639LP |
695/50 |
mPlum |
638nm |
FL10/LP7 |
639LP |
665/30 |
APC, Cy5, Alexa Fluor 647 |
FL11/LP6 |
685LP |
720/60 |
APC-Cy5.5, Alexa Fluor 700 |
FL12/LP5 |
752LP |
785/60 |
APC-Cy7, APC-Alexa Fluor 750 |
3. APPLICATIONS
Flow cytometers have the ability to measure multiple celular parameters and markers of a large number of cells at a rapid rate. So the applications of flow cytometry are numerous, and this has led to the widespread use of these instruments in biological research.
The main applications that we have developed in the center are:
• Cell viability assays
• Membrane potencial
• Inmunophenotyping
• Cell cycle analysis
• Apoptosis and necrosis
• Cell sorting
- Stem cells (mouse)
- Cell lines
- Drosophila discs cells (eyes and wings)
- Hematopoietic cells (mouse blood, kidney, liver, bone marrow, spleen, thymus)
4. FACILITY
4.1. Contact
Contact with
Corín Díaz Ramos
Telephone: 954 977432
E-mail: cdiaramwfxqx@qfwcpupo.es
4.2. Sample preparation
Samples must be prepared on aseptic conditions and must be filtered with a 40µm strainer. We recommend discussing previously the experimental design with the facility's staff.
4.3. Service
- Assisting researchers in designing experiments
- Analytical cytometers' user training
- Assisting researchers with analysis and data interpretation
- Sorting cell populations
5. BIBLIOGRAPHY
1. Flow cytometry and cell sorting (1992) A. Radbruch
Springer-Verlag, Berlin.
(579 FLO)
2. In living color: protocols in flow cytometry Rochelle A. Diamond, Susan DeMaggio
(579 IN)
3. Flow Cytometry. A Practical Approach (2000) Ormerod, MG.
3rd edition. Oxford University Press, Oxford. Robinson, J.P.,
(576 FLO)
4. Practical Flow Cytometry (2003) Howard Shapiro.
Fourth Edition Wiley-Liss, New York
(576 SHA)
5. Cytometry: new developments (2004)
San Diego: Elsevier
(576 CYT)
6. Flow cytometry protocols (2004)
Totowa, New Jersey, Humana Press, cop.
(576 HAW flo)
7. Flow cytometry: principles and applications (2007)
Totowa, New Jersey, Humana Press, cop.
(576 FLO)
6. EDUCATIONAL WEB PAGES