# Methods to determine the size of an object in microns

(Difference between revisions)
 Revision as of 12:42, 11 August 2005 (view source)← Previous diff Revision as of 12:54, 11 August 2005 (view source)Next diff → Line 5: Line 5: $\mbox{Pixel size} = \frac{6.45um}{\mbox{total magnification}}$ $\mbox{Pixel size} = \frac{6.45um}{\mbox{total magnification}}$ - Assuming that the magnifier is at position 1x (not 1.5x), this formula gives us: + This formula gives us: *60x  :  0.1075 um/pixel  or  9.30 pixels/um *60x  :  0.1075 um/pixel  or  9.30 pixels/um Line 11: Line 11: *20x  :  0.3225 um/pixel  or  3.10 pixels/um *20x  :  0.3225 um/pixel  or  3.10 pixels/um *10x  :  0.645  um/pixel  or  1.55 pixel/um *10x  :  0.645  um/pixel  or  1.55 pixel/um + + Important notes: + * We assumed here that the magnifier is at position 1x (not 1.5x). If it is at 1.5x, total magnification must be multiplied by 1.5. + * We assumed here that the bin size is 1x1. If it is, say, 2x2, the size of a pixel will be double. + + ===Calibration slide=== + + Use a calibration slide which has a grid with known line-to-line spacing. I borrowed such a slide from Peter Sorger's lab. + Results will be posted here soon. + + ===xy-motorized stage=== + + Put a sample on a slide or pad (grain of dust, cells, sphere). Record the position of the sample. Use IPLab to tell the stage to move a certain distance in um. Determine the distance (in pixels) between the sample's former position and its new position.

## Revision as of 12:54, 11 August 2005

### Mathematically

Pixels on our CCD are 6.45um. The size (in microns) of one pixel in an image will depend on how much the image was magnified before reaching the CCD:

$\mbox{Pixel size} = \frac{6.45um}{\mbox{total magnification}}$

This formula gives us:

• 60x  : 0.1075 um/pixel or 9.30 pixels/um
• 40x  : 0.1613 um/pixel or 6.21 pixels/um
• 20x  : 0.3225 um/pixel or 3.10 pixels/um
• 10x  : 0.645 um/pixel or 1.55 pixel/um

Important notes:

• We assumed here that the magnifier is at position 1x (not 1.5x). If it is at 1.5x, total magnification must be multiplied by 1.5.
• We assumed here that the bin size is 1x1. If it is, say, 2x2, the size of a pixel will be double.

### Calibration slide

Use a calibration slide which has a grid with known line-to-line spacing. I borrowed such a slide from Peter Sorger's lab. Results will be posted here soon.

### xy-motorized stage

Put a sample on a slide or pad (grain of dust, cells, sphere). Record the position of the sample. Use IPLab to tell the stage to move a certain distance in um. Determine the distance (in pixels) between the sample's former position and its new position.