Difference between revisions of "Final Project -- Nathan S Lachenmyer"

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I have two project proposals that I am deciding between:
 
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=Confocal Microscopy=
 
=Confocal Microscopy=
 
==Literature==
 
==Literature==
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* Handbook of biological confocal microscopy / editor, James B. Pawley.
 
* Handbook of biological confocal microscopy / editor, James B. Pawley.
  
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==Goal==
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==Goals and Checkpoints==
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* Minimum: Create a z-stack of images of a 3D sample
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===Goals===
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* Potential Improvements:
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{| border = "1" align="center" style="color: black; background-color: #ffffcc;" width="85%" class="wikitable"
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** 3D Reconstruction in Software
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!Key Factors!!Proposed Technical Targets!!Requirements for Success!!Current Practice!!Associated Technical Barriers
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** Laser Scanning for CLSM
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|- align="center"
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|style="width: 20%;"|OCLSM Cost
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|style="width: 20%;"|< $8,000
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|style="width: 20%;"|< $15,000
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|style="width: 20%;"|> $100,000 commercial; ~$30,000 DIY
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|style="width: 20%;"|Cheap replacement for PMT & Galvo, etc.
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|-
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|style="width: 20%;"|OCLSM Scan Time
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|style="width: 20%;"|1 frame/second
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|style="width: 20%;"|1 frame / 10 seconds
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|style="width: 20%;"|10-50 fps
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|style="width: 20%;"|Fast scanning mirrors + ADC time
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|}
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===Timeline===
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* 2 Weeks: Brightfield Confocal Microscopy
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* 2 Weeks: Software Image Capture, PSF Characterized, Z-Stack Images
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* 3 Weeks: Laser Scanning Features
  
 
==Proposed Design==
 
==Proposed Design==
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* Aperture to block out non-convergent rays
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*
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==Challenges==
 
==Challenges==
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* Control software
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* Getting good resolution
  
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=Ca<math>^{2+}</math> Imaging=
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=Weekly Updates=
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==Literature==
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==2012 April 8==
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* Imaging in Neuroscience and Development: A Laboratory Manual / editors, Rafael Yuste and Arthur Konnerth
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* Look for PMTs and/or Galvos
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* Handbook of biological confocal microscopy / editor, James B. Pawley.
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* Detection and Measurement of Free Ca2+ in Cells / editor, Ashley Campbell
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* Calcium Measurement Methods / editors, Alexei Verkhratsky and Ole Peterson
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==Goals==
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==2012 April 15==
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* Determine Ca<math>^{2+}</math> concentration in a biological system
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* Calculations for stuff
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* Potential Improvements:
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** Lifetime vs. Intensity Measurements
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** FRET Measurements
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==Proposed Design==
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==2012 April 22==
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* Fluorescence Microscope
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* Construct Galvos, Program Maple
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** 422 nm (brightfield, optical)
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** 340, 380 nm (fluorescence; two wavelengths provides ratiometric determination of [Ca])
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==2012 April 29==
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* Motorized Mirror or Filter (Servo) to switch wavelengths
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* Write-up for Fluorescence Microscope
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* Sync motor signal with camera signal (take pictures)
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* Post-Processing Analysis to determine [Ca](t)
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==Challenges=
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==2012 May 06==
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* Syncing up motor with camera for fluorescence imaging
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*

Latest revision as of 06:53, 7 May 2012

Confocal Microscopy

Literature

  • Imaging in Neuroscience and Development: A Laboratory Manual / editors, Rafael Yuste and Arthur Konnerth
  • Confocal microscopy for biologists / Alan R. Hibbs.
  • Handbook of biological confocal microscopy / editor, James B. Pawley.

Goals and Checkpoints

Goals

Key Factors Proposed Technical Targets Requirements for Success Current Practice Associated Technical Barriers
OCLSM Cost < $8,000 < $15,000 > $100,000 commercial; ~$30,000 DIY Cheap replacement for PMT & Galvo, etc.
OCLSM Scan Time 1 frame/second 1 frame / 10 seconds 10-50 fps Fast scanning mirrors + ADC time

Timeline

  • 2 Weeks: Brightfield Confocal Microscopy
  • 2 Weeks: Software Image Capture, PSF Characterized, Z-Stack Images
  • 3 Weeks: Laser Scanning Features

Proposed Design

Challenges

  • Control software
  • Getting good resolution

Weekly Updates

2012 April 8

  • Look for PMTs and/or Galvos

2012 April 15

  • Calculations for stuff

2012 April 22

  • Construct Galvos, Program Maple

2012 April 29

  • Write-up for Fluorescence Microscope

2012 May 06

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