Optical configuration of the FACSVantage DiVa.
In addition to the Digital Vantage (DiVa) upgrade, the FACSVantage has been upgraded with an 11-color optical bench. This bench upgrade and the DiVa electronics allow six-color analysis off the primary argon-ion 488 nm laser, three colors off the secondary HeNe 633 nm laser, and two colors off the krypton-ion UV or violet laser. In addition to being able to analyze greater numbers of fluorochromes simultaneously, the new bench gives us much greater flexibility for analyzing newly available fluorescent probes, such as some of the newer PE and APC tandem conjugates. The DiVa softare gives us simultaneous analysis off all fluorescent channels, as well as software-based compensation between all parameters.
(Below). Upgraded 11-color FACSVantage DiVa optical bench.
(Below). Upgraded 11-color FACSVantage DiVa optical bench. Primary (blue), secondary (red) and tertiary (violet) detectors are indicated.
(Below). Standard FACSVantage DiVa dichroics. These optics are the default set for the 11-color optical bench upgrade. Our older filter sets can be substituted where necessary.
| Dichroic | Use |
| 485 LP | separation of Cascade Blue from Cascade Yellow (or ELF-97) |
| 560 SP | separation of FITC and PE |
| 600 LP | separation of PE-Texas Red from FITC and PE |
| 640 SP | separation of PE-Cy5 from FITC, PE and PE-Texas Red |
| 685 LP | separation of APC from APC-Cy5.5 and APC-Cy7 |
| 740 LP | separation of PE-Cy5.5 from PE-Cy7, and APC-Cy5.5 from APC-Cy7 |
(Below). Standard FACSVantage DiVa filters.
| Filter | Use |
| 440/40 | Cascade Blue, Pacific Blue, DAPI, Hoechst 33258 and 33342 |
| 530/30 | FITC, Alexa Fluor 488 |
| 545/80 | Cascade Yellow, ELF-97, Yellow Fluorescent Protein |
| 575/42 | PE, PI |
| 610/20 | PE-Texas Red, DsRed |
| 660/20 | APC, Alexa Fluor 633 |
| 667/30 | PE-Cy5, Cy5, Alexa Fluor 647 |
| 710/50 | PE-Cy5.5, APC-Cy5.5, PE-Alexa Fluor 680, APC-Alexa Fluor 680, APC-Alexa Fluor 700 |
| 780/60 | PE-Cy7, APC-Cy7, PE-Alexa Fluor 750, APC-Alexa Fluor 750 |
Primary laser detectors. The primary argon-ion 488 nm laser now has six detectors (up from the original four on the FACSVantage SE). PE-Texas Red and PE-Cy5.5 can now be accomodated, allowing up to six-color analysis off the 488 nm laser line.
(Below). Primary laser detectors. Filter and dichroics are indicated.
Secondary laser detectors. The secondary HeNe 633 nm laser now has three detectors (up from the original two on the FACSVantage SE). APC, APC-Cy5.5 and APC-Cy7 (or the Alexa Fluor tandem equivalents) can be analyzed simultaneously. A fourth detector can be added for Texas Red incorporation into the above three-color scheme, if a yellow laser source is substituted for the red HeNe.
(Below). Secondary laser detectors. Filter and dichroics are indicated.
Tertiary laser detectors. The tertiary krypton-ion laser still has two detectors, as with the original FACSVantage SE optical bench. Ratiometric analysis of indo-1 for calcium concentration determination can be doine with the krypton-ion lase tuned to the UV, or Cascade Blue and ELF-97 with the laser tuned to the violet. The FL11 detector without intervening dichroic can be used for DAPI or Hoechst dye DNA content analysis.
(Below). Tertiary laser detectors. Filter and dichroics are indicated.
