Cell size measurements using light in flow cytometry and cell sorting

A new and improved flow cytometer means for measuring the size of a biological cell, comprising:

• 1. Means for directing said biological cells in column flow at a constant velocity;
• 2. Means for passing a laser beam having a cross section larger than each of said cells through said biological cells in column flow so that only one biological cell passes through said laser beam at a time;
• 3. Means for measuring the time of flight of each said biological cell through said laser beam deriving an electrical signal commensurate with the width and/or volume of each said biological cell or its nucleus, comprising:
• a. means for deriving a first electrical signal from the low angle forward light scatter characteristic or secondary fluorescent emission from each cell as it passes through the said laser beam;
• b. means for deriving a second electrical signal having an amplitude determined by the time integral of said first electrical signal;
• c. means for deriving a third electrical signal having an amplitude equal to the peak amplitude of said first electrical signal;
• d. means for deriving a fourth electrical signal by dividing the said second electrical signal by the said third electrical signal therefore deriving an electrical signal commensurate with the width of each said biological cell plus the width of the laser beam;
• e. means for deriving a fifth electrical signal by subtracting from said fourth electrical signal a constant electrical signal proportional to the width of the laser beam whereby the fifth electrical signal is directly proportional to the width of said biological cell.

A new and improved flow cytometer means for measuring the size of a biological cell, comprising:

• 1. Means for directing said biological cells in column flow at a constant velocity;
• 2. Means for passing a laser beam having a cross section larger than each of said cells through said biological cells in column flow so that only one biological cell passes through said laser beam at a time;
• 3. Means for measuring the time of flight of each said biological cell through said laser beam deriving an electrical signal commensurate with the width and/or volume of each said biological cell or its nucleus, comprising:
• a. means for deriving a first electrical signal from the secondary fluorescent emission from each cell as it passes through the said laser beam;
• b. means for deriving a second electrical signal having an amplitude determined by the time integral of said first electrical signal;
• c. means for deriving a third electrical signal having an amplitude equal to the peak amplitude of said first electrical signal;
• d. means for deriving a fourth electrical signal by dividing the said second electrical signal by the said third electrical signal therefore deriving an electrical signal commensurate with the width of each biological cell plus the width of the laser beam;
• e. means for deriving a fifth electrical signal by subtracting from said fourth electrical signal a constant electrical signal proportional to the width of the laser beam whereby the fifth electrical signal is directly proportional to the size of nucleus or cytoplasm of said biological cell.

A new and improved flow cytometer means for measuring the size of a biological cell, comprising:

• 1. Means for directing said biological cells in column flow at a constant velocity;
• 2. Means for passing a laser beam having a cross section larger than each of said cells through said biological cells in column flow so that only one biological cell passes through said laser beam at a time;
• 3. Means for measuring the time of flight of each said biological cell through said laser beam deriving an electrical signal commensurate with the width and/or volume of each said biological cell, comprising:
• a. means for deriving a first electrical signal from the low angle forward light scatter characteristic from each cell as it passes through the said laser beam;
• b. means for deriving a second electrical signal having an amplitude determined by the time integral of said first electrical signal;
• c. means for deriving a third electrical signal having an amplitude equal to the peak amplitude of said first electrical signal;
• d. means for deriving a fourth electrical signal by dividing the said second electrical signal by the said third electrical signal therefore deriving an electrical signal commensurate with the width of each said biological cell plus the width of the laser beam;
• e. means for deriving a fifth electrical signal by subtracting from said fourth electrical signal a constant electrical signal proportional to the width of the laser beam whereby the fifth electrical signal is directly proportional to the width of said biological cell.