ImagePro>Calibration problems

[<sabo_edmond[at]clalit.org.il>]

Dear image pro plus users,

I would like your advice about the following problem:

I am using the image pro plus version 4.5 for image analysis. I calibrated my system using the following method: 
I loaded a microscopic image of a grid captured at a certain magnification (for example x 40 magnifying lens), and I opened the calibration window in the IPP software. I opened the “Spacial”  window, I introduced “ microns” as measuring units, I pressed the first “image” button and I got the calibrating “line” on the screen. I indicated how many divisions on the grid correspond to a certain number of microns. I accepted the aspect ratio of 1 and I pushed the ok button. 
I considered my system to be calibrated for the corresponding magnification lens. Then, I started to measure cells and other microscopic objects.
Here is my problem: the system accurately measures the raw diameters of my objects (such as the Ferets, the longest and shortest diameters and so on). I know this for fact, since I validated these values using a different  image analysis software from a friend. 
When computing the objects area with the IPP software, my measurements were accurate only when the objects are square (rectangular) shaped. 
However, for any other shapes of the objects, the image pro plus system provides values of the area that are extremely large (two or three times) as compared to the communicated values obtained by other image analysis researchers.
It could be that I am doing something wrong when calibrating the system. I don’t think that there is a bug in the program (for calculating the area of an object).
I appreciate your help.
Yours,
Edmond

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