FCS Express: Kinetics Analysis

Most flow cytometers are capable of providing kinetics data by recording a time parameter in the FCS file. Kinetics experiments are particularly useful for examining changes in DNA content, antibody binding, and calcium flux. By adding a fluorescent marker to a sample and monitoring influx, efflux, or binding of the dye over time, unique information can be gathered. FCS Express offers the capability to display and analyze kinetics data in an easy and user friendly manner while updating your plots as your analysis changes.

FCS Express can plot many different parameters and statistics vs time such as:

  • Arithmetic Mean
  • Geometric Mean
  • Median
  • Count
  • Percentage Above Threshold
  • Any Parameter Math functions/formulas

 

Kinetics Plots Function Like Any Other Plot in FCS Express

FCS Express allows the user to format and gate any plot, show statistics, insert tokens, and does so in real time so your analysis updates as changes are made to your layout.

Watch a short webinar about how to perform Kinetics analysis in FCS Express

 

Using Kinetics for Quality Control

Kinetics Plots allow you to monitor the number of events/counts over time enabling the user to look for interruptions or peaks in the flow rate.

Sample #1 Gated on Lymphocytes
Sample #1 displaying counts over time. Note the valley where the sample was removed, drug added,and place back on the cytometer. The peak after the valley indicates the sample rate increased for a brief period of time.
Sample #1 displaying counts over time in logarithmic scale to minimize "noise".

 

Using Parameter Math to Set-Up Kinetics Ratios

Often times it is useful to define a ratio of one dye to another dye when examining kinetics data. The parameter math feature of FCS Express allows the user to define mathematical operations from simple ratios to complex formulas for use in your analysis. FCS Express will also calculate and display the actual ratio values rather than a multiplied integer as some other software packages do.

Setting up a ratio of FL1/FL3 to be used in analysis.
FL-1 vs Time
FL-3 vs Time
Ratio of FL-1/FL-3 created by parameter math allows further discrimination of the sample

 

Using Overlays with Kinetics Plots

Just like any other 1D Plot in FCS Express, Kinetics Plots can be overlayed to view multiple samples at once.

 

Calculating % Above Threshold

With FCS Express you can set gates for a baseline (Threshold) value and post treatment value and plot the % Above Threshold value. Notice in the plots below how the raw data from FL-1 (Top Row) and a Ratio (Bottom Row) was converted to a density plot, gated, and replotted as % Above Threshold.

Click on any plot to enlarge the image
Fl-1 vs Time - Kinetics Plot Raw Data FL-1 vs Time - Density Plot, Gated for baseline and Post Drug Treatment % Post Drug Treatment Above Baseline Gate vs Time
FL-1/FL-3 Ratio (Ca++ Ratio) vs Time - Kinetics Plot Raw Data FL-1/FL-3 Ratio (Ca++ Ratio) - Density Plot, Gated for baseline and Post Drug Treatment % Post Drug Treatment Above Baseline Gate vs Time