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Home > Imaging Software > Plug-In Module > 3D Constructor
3D Constructor Sharp Stack AFA Deconvolution
 
 
 
 
- Overview
- Advantages over Confocal Microscopy
- SharpStack System Requirements
- Elucidating 3D Structure
- Digital deconvolution
 

One frame with an AOI selected in a Z-stack on the left with two AOIs
and their deconvolved counterparts;
de-focused light contribution from other planes has been eliminated
For Image-Pro Plus 4.5.

 


Z-stack image·ÎºÎÅÍ ¼±¸íÇÑ Image¸¦ ÃßÃâÇϱâ À§Çؼ­´Â Ưº°ÇÑ ±â¼úÀÌ ¿ä±¸µË´Ï´Ù.
Image Pro PlusÀÇ Ãß°¡µÈ ModuleÀÎ SharpStackÀ» ÀÌ¿ëÇÏ¿© º¸´Ù ¼±¸íÇÑ Image¸¦ ¾òÀ¸½Ç ¼ö ÀÖ½À´Ï´Ù.
SharpStack´Â Image Analysis SoftwareÀÎ Image Pro Plus¿Í ÇÔ²² Image capture, 2D processing, ºÐ¼®°ú º¸°í¼­ ÀÛ¼ºµîÀ» ¼Õ½±°Ô ÇÒ ¼ö ÀÖ½À´Ï´Ù.
SharpStack¿¡¼­´Â Nearest neighbor, No neighbor, Inverse algorithm ±â´ÉÀ» »ç¿ëÇÏ¿© Çϳª ¶Ç´Â ¿©·¯ °³ÀÇ Z-Stack Image¸¦ ¼Õ½¬¿î ¹æ¹ýÀ¸·Î ¼±¸íÇÏ°Ô ¸¸µé ¼ö ÀÖ½À´Ï´Ù.

 

¼¼Æ÷¿Í Á÷¹°Àº 3Â÷¿ø ±¸Á¶ÀÔ´Ï´Ù. µû¶ó¼­ Çö¹Ì°æ¿¡¼­ °üÂû ÇÒ °æ¿ì Ç¥º»ÀÇ ÇÑ ºÎºÐ¿¡ focus°¡ ¸ÂÀ»Áö¶óµµ ´Ù¸¥ ºÎºÐÀÇ focus°¡ ¸ÂÁö ¾Ê½À´Ï´Ù. ÀÌ·¯ÇÑ ÀÌÀ¯·Î ±¤ÇÐÀûÀÎ Image¿¡¼­ÀÇ »ý¹°ÇРǥº» 3D Çؼ®Àº ´Ù¼Ò Á¦ÇÑÀÌ µË´Ï´Ù. ÀÌ·¯ÇÑ ¹®Á¦¿Í 3D dataÀÇ ³ôÀº ½Å·Ú¼ºÀ» À§Çؼ­ Confocal- Microscopy, Digital DeconvolutionÀ» »ç¿ëÇÏ½Ç ¼ö ÀÖ½À´Ï´Ù.
 

Confocal microscope´Â Photomultiplier tube (PMT)·Î µµ´ÞÇÏ´Â Èñ¹ÌÇÑ ¿µ»ó (focus°¡ ¸ÂÁö ¾Ê´Â)À» Á¦ÇÑÇϱâ À§ÇØ Pinhole aperture¸¦ »ç¿ëÇÕ´Ï´Ù.
Pinhole aperture¹æ½ÄÀ¸·Î ÀÎÇØ ´ë¹°·»Á ÅëÇؼ­ ¼öÁýµÈ ¸ðµç Çü±¤½ÅÈ£°¡ ¸ðµÎ »ç¿ëµÇÁö ¸øÇϹǷΠÀÌ ½ÅÈ£¸¦ ÁõÆø ½ÃÅ°±â À§ÇØ Áõ°­µÈ excitation laser intensity¸¦ »ç¿ëÇÏÁö¸¸, Á¾Á¾ Image°¡ ¼Õ»óµÇ°Å³ª ¹éÈ­Çö»óÀÌ ¹ßÇöÇÒ °¡´É¼ºÀÌ ÀÖÀ¸¸ç ¼Õ»óÀ» ÁÙ ¼ö ÀÖ½À´Ï´Ù.
¶ÇÇÑ ´Ù¸¥ Microscope¿¡ ºñÇØ »ó´ëÀûÀ¸·Î °í°¡ÀÔ´Ï´Ù.
 

Confocal System °ú´Â ´Þ¸®, Digital Deconvolution Microscopy´Â ´ë¹°·»Áî¿¡ ÀÇÇØ ¼öÁýµÈ ¸ðµç Çü±¤½ÅÈ£¸¦ Pinhole¹æ½ÄÀ» »ç¿ëÄ¡ ¾Ê°í ¹Ù·Î °í°¨µµ CCD Ä«¸Þ¶ó¿¡¼­ ¹Þ¾Æ µéÀÏ ¼ö ÀÖ½À´Ï´Ù.  Image SystemÀÇ PointSpread Function(PSF)¿¡ ÀÇÇØ Digital Deconvolution´Â Image QualityÀÇ ÀúÇϸ¦ ÀúÀåÇϸç, Focus°¡ ¸ÂÁö ¾Ê´Â Èñ¹ÌÇÑ ¿µ»óÀº °¢±â ´Ù¸¥ ±¤ÇÐÀÇ
¿µ¿ª¿¡¼­ ImageÈ­ µË´Ï´Ù.
Linear, shift-invariant systemÀ¸·Î¼­ ±¤ÇÐÇö¹Ì°æ»óÀ» modelingÇÒ °æ¿ì, PSF´Â Çö¹Ì°æ¿¡¼­ÀÇ ¾î¶°ÇÑ Imageº¯Çüµµ ³ªÅ¸³¾ ¼ö ÀÖ½À´Ï´Ù. ÀüÇüÀûÀÎ Çü±¤ Çö¹Ì°æ Image´Â ´ÙÀ½°ú °°ÀÌ Ç¥ÇöµÉ ¼ö ÀÖ½À´Ï´Ù.
[measured image] = [PSF] * [desired image]
¿©±â¼­ '*'Ç¥½Ã´Â ConvolutionÀÇ ¼öÇÐÀû ¿¬»êÀÚ¸¦ ³ªÅ¸³À´Ï´Ù.

Deconvolution (*-1), ConvolutionÀÇ ¼öÇÐÀû ¿ª¼ö´Â ¾Æ·¡¿Í °°½À´Ï´Ù.
[desired image] = [measured image] (*-1) [PSF]
DeconvolutionÀÇ ¸ñÀûÀº ¿øÇÏ´Â Image¸¦ À§ÇÑ ¹æÁ¤½ÄÀ» °è»êÇÏ´Â °ÍÀÔ´Ï´Ù.
Inverse filtering ÀÌ·ÐÀ» ¹ÙÅÁÀ¸·Î ÇÑ Inverse Filter´Â ¹Ýº¹ °úÁ¤ÀÌ ¾ø°í, ´Ü°èÀûÀÔ´Ï´Ù.
Point Spread Function(PSF)°ú ÇÔ²² ImageÀÇ ConvolutionÀ¸·Î ¼±¸íÇÏÁö ¾ÊÀº Image¸¦ ³ªÅ¸³¾ ¼ö ÀÖ½À´Ï´Ù.
ÀϹÝÀûÀÎ ConvolutionÀº Optical transfer ±â´É°ú ÇÔ²² sampleÀÇ Fourierº¯È¯ÀÇ °öÀ¸·Î º¯È¯µË´Ï´Ù. Optical transfer function (OTF)Àº Point Spread Function (PSF)ÀÇ Fourier º¯È¯ÀÔ´Ï´Ù. Inverse Filter¸¦ Àû¿ëÇϸé OTF¸¦ ÀÌ¿ëÇÑ ImageÀÇ Fourierº¯È¯À» ºÐÇÒÇÏ¿© Image¸¦ ¿ø»óÅ·Πȸº¹ ½Ãų ¼ö ÀÖ½À´Ï´Ù.

Nearest Neighbor algorithmÀº Çѹø¿¡ ÇϳªÀÇ Image´Ü¸éÀ» ´ë»óÀ¸·Î Deconvolving(¹øÁüÁ¦°Å)ÀÛ¾÷À» ÇÒ ¼ö ÀÖ½À´Ï´Ù.
ÀÛ¾÷ÀÌ ÁøÇàµÇ´Â °úÁ¤¿¡¼­ Image ´Ü¸éÀ¸·ÎºÎÅÍÀÇ Á¤º¸´Â Image´Ü¸éÀÇ »óºÎ¿Í ÇϺο¡ ÀÖ´Â ÀÎÁ¢µÈ À̹ÌÁö°¡ »ç¿ëµË´Ï´Ù.
ÃøÁ¤µÈ ´Ü¸éÀ¸·ÎºÎÅÍÀÇ Á¤È®ÇÑ ´Ü¸é À§Ä¡ÀÇ Áõ°¡´Â »ç¿ëÀÚ°¡ ¼±ÅÃÇÒ ¼ö ÀÖ½À´Ï´Ù.
Á¤È®ÇÑ ´Ü¸éÀ» ¼±ÅÃÇÏ°Ô µÇ¸é ±× °á°ú´Â inverse filter ¹æ¹ý°ú °ÅÀÇ ±Ù»çÇÏ°Ô ³ª¿ÀÁö¸¸ inverse filterº¸´Ù Á»´õ ªÀº ½Ã°£¿¡ ÇÒ ¼ö ÀÖ½À´Ï´Ù. No Neighbor ¹æ¹ýÀº 2D PSF¸¦ ±¸¼ºÇϱâ À§ÇØ °¢°¢ÀÇ ´ÜÀÏ ´Ü¸éÀ¸·ÎºÎÅÍ µé¾î¿À´Â Á¤º¸°¡ »ç¿ëµË´Ï´Ù. ÀÌ ¹æ¹ýÀº ªÀº ½Ã°£ ¾È¿¡ ó¸®µÉ ¼ö ÀÖÁö¸¸ ´ÜÁ¡À̶ó ÇÏ¸é ´Ù¸¥ ¹æ¹ý¿¡ ºñÇØ ÁÁÀº °á°ú¸¦ ±â´ëÇÒ ¼ö´Â ¾ø½À´Ï´Ù.


Figure 3: Example of spherically aberrated image: a) XY and b) XZ maximum intensity projection of the original image*, c) XY and d) XZ maximum intensity projection of the deconvolved image with spherical aberration compensation (algorithm provided by AutoQuant Imaging, Inc.).

 


* Pig cerebellum image stack deconvoled and rendered with SharpStack and 3D Constructor plug-ins modules


- ImagePro-Plus Ver 4.5 ÀÌ»ó
- Pentium¥² 450MHz ÀÌ»ó.
- 256Mb Ram ÀÌ»ó(512Mb ±ÇÀå)
- Color monitor 16bit high color(24bit ¶Ç´Â 32bit ±ÇÀå)
- Çϵå¿þ¾î °¡¼Ó±â¸¦ Æ÷ÇÔÇÑ OpenGL °¡¼Ó ±×·¡ÇÈ Ä«µå(32Mb)

 
 
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