Paul Matsudaira WI/MIT BioImaging Center, Dept Biology and Div Biological Engineering, MIT Cellular machinery, biomechanics, and bioinformatics IC-21 macrophage cells transfected with GFP, imaged in 3D, and rendered as a projected solid (Imaris) bioengineering models describe biological processes complex movements via cellular machines mechanics + chemistry structures capture states of movement informatics/computing resources

Quantitative Biology Paradigm courtesy of CSBi

Cell biologists study large, dynamic structures G. Borisy and S. Tatiana

1 nm1 µm1 mm cellsorganelles molecules protein machines NMR, x-rays cryo electron microscopy single particle det. dynamic 4D light microscopy resolution and size scale Overlapped structural and cell biology approaches

zone of adhesion and extenstion Forces generate movements, machines generate forces fimbrin-EGFP timelapse 30 sec

Changes in structure-assembly, force, chemistry mother adhesion daughter adhesions topside Formation of cell adhesions at the leading edge of a macrophage cell (J. Evans et al J. Cell Biol. in press)

multispectral fluorescence data acquisition video rate data acquisition cellular tomography and single particle cryoEM imaging high content screening whole proteome studies Cell Biology Appetite for More Information

High content: Tracking all cell adhesions (J. Evans et al J. Cell Biol. in press)

Dynamics of structure modulated by microtubules 10 µm paclitaxel 10 µm demecolcine (J. Evans et al J. Cell Biol. in press)

Imaging storage/processing requirements objectfile size genome sequence 12 MB protein structures 1 2 GB 2D localization 2 6 GB 3D localization GB 4D localization TB 1 52kDa (ave. MW) x 110 MW/residues x 10 atoms 2 512x512 two channel, 16 bit TIFF 3 50 image slices/stack 4 12 images/series

The BioImaging Pipeline acquisition management processing analysis modelling

Podosomes split, merge, and appear de novo polar assembly simple dendroid

genomicsimaging tracescounts ATGCvoxel sequenceimagefunction Imaging is an informatics science

WI/MIT BioImaging Net terminal Origin TB confocal microscopy Origin TB IBM Power Cryo-EM 2-photon microscopy Tape RAID 30TB imaging modes

Expansion and management of imaging data channel 0channel 1channel 2 multi-spectral (channels) 12 or 16-bit acquisition 256 x KB KB 1024x MB 3 14 MB image size (pixels)channels file size 1+2+3

(x,y,z,t,ch) file size 256 x 256 x 1 x KB 256 x 256 x 50 x 3 20 MB 256 x 256 x 50 x 20 x MB 1024 x 1024 x 50 x MB 1024 x 1024 x 50 x 20 x 3 16 GB 3D/4D image data (stacks) 35KB to 8.2 MB per channel/stack (X:Y:Z or X:Y:T)

NMR x-ray cryoEM light microscopy

raw data 12 or 16-bit deconvolved data 32-bit intermediates 32-bit raw data 32-bit 40 MB80 MB 40 MB120 MB200 MB280 MB360 MB accumulated file size Image processing (deconvolution) RAM 256 x 256 x 50 x 3 (x,y,z,ch)

3.2 GB ch1 vs. ch2 ch1 vs. ch3 ch2 vs. ch3 ch1 vs ch2 vs. ch3 colocalization Total data accumulation rendering 1 GB movies object analyses 1 GB 1 GB ? raw0.4 GB post-deconvol3.6 GB analysis7.2 GB total 11.2 GB

terminal SGI Origin GB RAM 4 TB Zeiss 510 META ftp data acquisition data storage image restoration image analysis batch rendering database management remote visualization rendering batch design Current WI (local) imaging net stack (512 x 512 x 50 x 12 x 6) 4.8 GB

Microtubules at podosomes tubulin actin fimbrin

Dynamic structures from light microscopy fimbrin-EGFP timelapse 15 sec side/bottom view top view ‘finger’ ‘palm’ ‘finger’ ‘finger-tip’ ‘palm’ assembly of cell adhesions in a live cell

Dynamic Cell Adhesions - Static