Difference: PPFiducialsTomography (1 vs. 16)

Revision 1621 Sep 2009 - Main.KdDerr

 
META TOPICPARENT name="CemProtocGrids"
Return to Cryo EM web page at http://www.nysbc.org/facilities/CEM

Principles & Protocols

Changed:
<
<

Protocols - Grid preparation

>
>

Protocols - Fiducial Markers for Tomography

 

Jump to Principles

Deleted:
<
<

Application of Gold Beads to sections as Fiducial Markers for Tomography

 Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

META TOPICMOVED by="KakoliMitra" date="1234475869" from="Main.CemPPGridsFTO" to="Main.PPFiducialsTomography"

Revision 1521 Sep 2009 - Main.KdDerr

 
META TOPICPARENT name="CemProtocGrids"
Added:
>
>		Return to Cryo EM web page at http://www.nysbc.org/facilities/CEM
 
Changed:
<
<

Principles & Protocols

>
>

Principles & Protocols

Deleted:
<
<
 
Added:
>
>

Protocols - Grid preparation

Jump to Principles

 

Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

META TOPICMOVED by="KakoliMitra" date="1234475869" from="Main.CemPPGridsFTO" to="Main.PPFiducialsTomography"

Revision 1419 Aug 2009 - Main.KdDerr

 
META TOPICPARENT name="CemProtocGrids"
Deleted:
<
<		 NYSBC|Cryo-EM P&P: Grid preparation - Fiducials for tomography
 
Changed:
<
<		Principles & Protocols
>
>

Principles & Protocols

Added:
>
>
 
Changed:
<
<
>
>

Application of Gold Beads to sections as Fiducial Markers for Tomography

Deleted:
<
<
Interested in EM? Using CEMfac? Principles & Protocols CEMfac Equipment Seminars & Courses
CEMfac Home? CEM Introduction Microscope Schedule NYSBC Directory Cryo-EM public website

Grid preparation

Protocols - Fiducials for tomography

Contents

Application of Gold Beads to sections as Fiducial Markers for Tomography
 Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

META TOPICMOVED by="KakoliMitra" date="1234475869" from="Main.CemPPGridsFTO" to="Main.PPFiducialsTomography"

Revision 1312 Feb 2009 - Main.KakoliMitra

 
META TOPICPARENT name="CemProtocGrids"
NYSBC|Cryo-EM P&P: Grid preparation - Fiducials for tomography

Principles & Protocols
Interested in EM? Using CEMfac? Principles & Protocols CEMfac Equipment Seminars & Courses
Changed:
<
<
CEMfac Home NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website
>
>
CEMfac Home? CEM Introduction Microscope Schedule NYSBC Directory Cryo-EM public website
 

Grid preparation

Protocols - Fiducials for tomography

Contents

Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

Added:
>
>
 
META TOPICMOVED by="KakoliMitra" date="1234475869" from="Main.CemPPGridsFTO" to="Main.PPFiducialsTomography"

Revision 1212 Feb 2009 - Main.KakoliMitra

 
META TOPICPARENT name="CemProtocGrids"
NYSBC|Cryo-EM P&P: Grid preparation - Fiducials for tomography

Principles & Protocols
Interested in EM? Using CEMfac? Principles & Protocols CEMfac Equipment Seminars & Courses
Changed:
<
<
CEMfac Home NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website
>
>
CEMfac Home NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website
 

Grid preparation

Protocols - Fiducials for tomography

Contents

Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

META TOPICMOVED by="KakoliMitra" date="1234475869" from="Main.CemPPGridsFTO" to="Main.PPFiducialsTomography"

Revision 1112 Feb 2009 - Main.KakoliMitra

 
META TOPICPARENT name="CemProtocGrids"
NYSBC|Cryo-EM P&P: Grid preparation - Fiducials for tomography

Principles & Protocols
Interested in EM? Using CEMfac? Principles & Protocols CEMfac Equipment Seminars & Courses
Changed:
<
<
CEMfac Home? NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website
>
>
CEMfac Home NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website
 

Grid preparation

Protocols - Fiducials for tomography

Contents

Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

META TOPICMOVED by="KakoliMitra" date="1234475869" from="Main.CemPPGridsFTO" to="Main.PPFiducialsTomography"

Revision 1012 Feb 2009 - Main.KakoliMitra

 
META TOPICPARENT name="CemProtocGrids"
NYSBC|Cryo-EM P&P: Grid preparation - Fiducials for tomography

Principles & Protocols
Interested in EM? Using CEMfac? Principles & Protocols CEMfac Equipment Seminars & Courses
CEMfac Home? NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website

Grid preparation

Protocols - Fiducials for tomography

Contents

Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

Changed:
<
<
META TOPICMOVED by="KakoliMitra" date="1234475288" from="Main.GoldBeads" to="Main.CemPPGridsFTO"
>
>
META TOPICMOVED by="KakoliMitra" date="1234475869" from="Main.CemPPGridsFTO" to="Main.PPFiducialsTomography"
 

Revision 912 Feb 2009 - Main.KakoliMitra

 
META TOPICPARENT name="CemProtocGrids"
NYSBC|Cryo-EM P&P: Grid preparation - Fiducials for tomography

Principles & Protocols
Interested in EM? Using CEMfac? Principles & Protocols CEMfac Equipment Seminars & Courses
CEMfac Home? NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website

Grid preparation

Protocols - Fiducials for tomography

Contents

Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
Changed:
<
<
  • Positive stain? sample grids
>
>
 
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

META TOPICMOVED by="KakoliMitra" date="1234475288" from="Main.GoldBeads" to="Main.CemPPGridsFTO"

Revision 812 Feb 2009 - Main.KakoliMitra

 
META TOPICPARENT name="CemProtocGrids"
NYSBC|Cryo-EM P&P: Grid preparation - Fiducials for tomography

Principles & Protocols
Interested in EM? Using CEMfac? Principles & Protocols CEMfac Equipment Seminars & Courses
CEMfac Home? NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website

Grid preparation

Protocols - Fiducials for tomography

Contents

Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain? sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

Added:
>
>
META TOPICMOVED by="KakoliMitra" date="1234475288" from="Main.GoldBeads" to="Main.CemPPGridsFTO"
 

Revision 712 Feb 2009 - Main.KakoliMitra

Changed:
<
<
META TOPICPARENT name="SamplePreparation"
>
>
META TOPICPARENT name="CemProtocGrids"
  NYSBC|Cryo-EM P&P: Grid preparation - Fiducials for tomography

Principles & Protocols
Interested in EM? Using CEMfac? Principles & Protocols CEMfac Equipment Seminars & Courses
CEMfac Home? NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website

Grid preparation

Protocols - Fiducials for tomography

Contents

Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain? sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

Revision 612 Feb 2009 - Main.KakoliMitra

 
META TOPICPARENT name="SamplePreparation"
NYSBC|Cryo-EM P&P: Grid preparation - Fiducials for tomography

Principles & Protocols
Interested in EM? Using CEMfac? Principles & Protocols CEMfac Equipment Seminars & Courses
CEMfac Home? NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website

Grid preparation

Protocols - Fiducials for tomography

Contents

Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain? sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry
Deleted:
<
<
 
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

Revision 512 Feb 2009 - Main.KakoliMitra

 
META TOPICPARENT name="SamplePreparation"
Changed:
<
<		Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain? sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry

  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Visit Cryo EM Web site at http://www.nysbc.org/facilities/CEM

-- DavidStokes - 18 Mar 2005

>
>		 NYSBC|Cryo-EM P&P: Grid preparation - Fiducials for tomography

Principles & Protocols
Interested in EM? Using CEMfac? Principles & Protocols CEMfac Equipment Seminars & Courses
CEMfac Home? NYSBC & Cryo-EM Information? Microscope Schedule NYSBC Directory Cryo-EM public website

Grid preparation

Protocols - Fiducials for tomography

Contents

Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
Added:
>
>
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain? sample grids
  • Sonicate gold solution 10-15 min
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
  • Allow to air dry

  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Public website mirror

-- DavidStokes - 18 Mar 2005

 

Revision 408 Jun 2005 - Main.KdDerr

 
META TOPICPARENT name="SamplePreparation"
Application of Gold Beads to sections as Fiducial Markers for Tomography

Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.

You will need:

  • 200 mesh grids with sections or a thin film
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
  • glow discharge

Procedure:

  • Put on gloves!
  • Positive stain? sample grids
  • Sonicate gold solution 10-15 min
Changed:
<
<
  • Glow Discharge the grid
  • Place grid sample side down on a drop of 5nm or 10 nm gold solution
  • Allow to air dry
>
>
  • Glow Discharge the grid with sections
  • Place grid sample side up on parafilm
  • Deposit 1um of 10 nm gold solution
Added:
>
>
  • Allow to air dry
 
  • Apply sample or view grid

Do not use copper grids for immunogold labeling

Visit Cryo EM Web site at http://www.nysbc.org/facilities/CEM

-- DavidStokes - 18 Mar 2005

Revision 322 Apr 2005 - Main.KdDerr

 
META TOPICPARENT name="SamplePreparation"
Application of Gold Beads to sections as Fiducial Markers for Tomography

Added:
>
>		Gold Fiducial markers can be applied before or after the sections are placed on the grid. If they are applied before the sections, a thin film must be present for the gold to adhere to.
 

You will need:

Changed:
<
<
  • 30-70 nm sections on grid
>
>
  • 200 mesh grids with sections or a thin film
Deleted:
<
<
  • 0.01% poly-L -lysine
 
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers
Added:
>
>
  • glow discharge
 
Deleted:
<
<
 

Procedure:

  • Put on gloves!
  • Positive stain? sample grids
  • Sonicate gold solution 10-15 min
Changed:
<
<
  • Float sample on a drop of 0.01% poly-L-lysine 7-10min
  • Rinse with DD H20
  • Air dry 2 min
  • Place grid sample side down on a drop of 5nm gold solution for 2min
  • DD H2O rinse
>
>
  • Glow Discharge the grid
  • Place grid sample side down on a drop of 5nm or 10 nm gold solution
  • Allow to air dry

  • Apply sample or view grid
Deleted:
<
<
  • Air dry 2 min
  • Place grid, sample side up on a drop of 10nm gold solution
  • DD H2O rinse
  • Air dry
 Do not use copper grids for immunogold labeling

Visit Cryo EM Web site at http://www.nysbc.org/facilities/CEM

-- DavidStokes - 18 Mar 2005

Revision 222 Mar 2005 - Main.DavidStokes

 
META TOPICPARENT name="SamplePreparation"
Application of Gold Beads to sections as Fiducial Markers for Tomography

You will need:

  • 30-70 nm sections on grid
  • 0.01% poly-L -lysine
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers

Procedure:

  • Put on gloves!
  • Positive stain? sample grids
  • Sonicate gold solution 10-15 min
  • Float sample on a drop of 0.01% poly-L-lysine 7-10min
  • Rinse with DD H20
  • Air dry 2 min
  • Place grid sample side down on a drop of 5nm gold solution for 2min
  • DD H2O rinse
  • Air dry 2 min
  • Place grid, sample side up on a drop of 10nm gold solution
  • DD H2O rinse
  • Air dry

Do not use copper grids for immunogold labeling

Visit Cryo EM Web site at http://www.nysbc.org/facilities/CEM

-- DavidStokes - 18 Mar 2005

Added:
>
>
 

Revision 118 Mar 2005 - Main.DavidStokes

 
META TOPICPARENT name="SamplePreparation"
Application of Gold Beads to sections as Fiducial Markers for Tomography

You will need:

  • 30-70 nm sections on grid
  • 0.01% poly-L -lysine
  • Parafilm
  • Sonicator
  • Colloidal gold solution 5 & 10nm
  • DD H2O
  • Filter paper
  • Tweezers

Procedure:

  • Put on gloves!
  • Positive stain? sample grids
  • Sonicate gold solution 10-15 min
  • Float sample on a drop of 0.01% poly-L-lysine 7-10min
  • Rinse with DD H20
  • Air dry 2 min
  • Place grid sample side down on a drop of 5nm gold solution for 2min
  • DD H2O rinse
  • Air dry 2 min
  • Place grid, sample side up on a drop of 10nm gold solution
  • DD H2O rinse
  • Air dry

Do not use copper grids for immunogold labeling

Visit Cryo EM Web site at http://www.nysbc.org/facilities/CEM

-- DavidStokes - 18 Mar 2005

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