MultiCycle AV for FCS Express is capable of autofitting the appropriate Cycle to suit your data.  However, users have the ability to override the autofit and choose a Cycle to fit to their data.

 

Here is a summary of the various Cycles that are available in MultiCycle AV.

 

Cycles

1 Cycle

The most common and frequently used.  Used when there is 1 full cell cycle present.

1 Cycle Standard Overlay Peak

Similar to 1 Cycle.  Ideal when using a standardization tool like chicken erythrocytes.

1 Cycle Apoptotic Overlay Peak

This cycle is performed when it is assumed that there is one extra peak to the left of the G0/G1 peak.

1 Cycle Aneuploid Overlay Peak

This cycle is performed when there is an Aneuploid peak present that does not contain a full cell cycle.

1 Cycle Diploid Overlay Peak

This cycle is performed when there is a Diploid peak that does not contain a full cell cycle.

2 Cycle

This cycle is used when there are 2 full cell cycles present.

3 Cycle

This cycle is used when there are 3 full cell cycles present.

Synchronous S Phase

This cycle should be used when performing an experiment where S phase is arrested, leading to a peak in S phase, rather than a smooth gradual curve.

 

 

MultiCycle AV for FCS Express applies six different models to your data at the same time.  

 

In FCS Express, you can create a DNA Model Statistics Summary box that shows your statistics as it applies to each of the six different models (Figure 5.14).

 

 

 

Figure 5.14 - DNA Model Statistics

Figure 5.14 - DNA Model Statistics

 

 

Here is a summary of the various Cycles that are available in MultiCycle AV.

 

Models

1) SL S0

Most common model used.  Sliced nuclei background modeling with zero order S phase, but without other options.  Used when debris curve looks like a ski slope.

2)SL CL S0

Sliced nuclei background modeling with clumping compensation with zero order S phase.  Used if debris curve is flat and cuts into G0/G1.

3)SL G21 S0

Sliced nuclei background modeling, plus the constraint of the G2/G1 ratio.  For 1 cycle fitting models, the G2/G1 ratio is the current default stored within Multicycle AV.

 

Please note, for 2 Cycle models, the G2/G1 ratio of the diploid cycle is constrained to be equal to the aneuploid G2/G1 ratio that resulted from the user-chosen fitting model.

 

For 3 Cycle models, the G2/G1 ratio of all 3 cell cycles is constrained to be equal to the average of the 2 aneuploid G2/G1 ratios that resulted from the user-chosen fitting model.

4)SL CL G21 S0

Sliced nuclei modeling plus aggregation modeling and addition of G2/G1 constraint, as detailed above.  Basically, this is a software version of doublet discrimination.  This model looks at all possible combinations of G1 and G2 stuck together and figures out the % of cells that are stuck together.

5)SL S1

A 1st order S phase polynomial is used with sliced nuclei background modeling.  This model is to be used when the S phase is more in the shape of a broadened trapezoid.

 

S order = 0 (broadened rectangle)

S order = 1 (broadened trapezoid)

S order = 2 (higher on the sides, lower in the middle)

6)SL CV S1

Sliced nuclei modeling with a 1st order S phase, plus the CVs of all peaks are constrained.  This model can be used when the G2/M peak is buried in the S phase.

 

 

 

References for this information can be found at our FCS Express - Multicycle knowledgebase page.

 

In cases where the data is suboptimal or the cell cycle peaks are not well defined, users can get Intra-model or Inter-model errors.  You can find the definition for these errors at the FCS Express - Multicycle knowledgebase page.

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