Transmission electron microscopy (TEM)

The mPrep System™ streamlines Transmission Electron Microscopy (TEM) sample preparation using a capsule based approach. Use mPrep/s capsules to fix, orient, embed, and section specimens. Use mPrep/g capsules to stain or immuno-label TEM grids.

The mPrep System efficiently produces quality results from every sample.  Imagine this in your lab.

  • As few as two human touches from microtome to microscope – reduces damage and loss
  • Grids and capsules labeled for easy tracking from start to storage
  • Capsules attach to common lab pipettors for controlled reagent timing and minimal reagent consumption
  • Parallel processing
    • Stain from one to dozens of grids simultaneously using multi-channel pipettors
    • Automate using the mPrep-ASP-1000 Automated Specimen Processor
    • Identical reagent timing
    • Reduced tedium and labor costs

 

The mPrep System is designed for versatility. Our customers continue to develop new applications. Explore some TEM applications below.

Kidney (TEM)

Nanoparticles (TEM negative stain)

TEM applications

Biological tissue fixation, embedding and orientation

The mPrep System streamlines the entire tissue preparation and grid staining workflow.

Problems

Tissue prep for TEM typically involves over 20 reagent exchanges per specimen, using laboratory equipment not well suited for this purpose.  This presents microscopists with multiple challenges:

  • Difficulty achieving uniform processing times across multiple samples
  • Reagent consumption excessive relative to the amount required to ensure reaction
  • Achieving proper orientation is time consuming
  • Specimens damage from contact with air, vials, and transfer pipettes
  • Lost specimens when difficult to see or they stick to containers

 

mPrep Solutions

  • Simultaneous processing of reagent exchanges assures uniform timing
  • Controlled reagent usage with pipettors = reduced reagent consumption
  • Accurate orientation in seconds using the mPrep Workstation
  • Encapsulated specimen reduces handling = fewer damaged specimens
  • Entrapped specimens are harder to lose. Will not float.
  • Labeled capsules are easy to track.

Heart (TEM)

Skin (TEM)

Application Notes

This application note illustrates mammalian kidney tissue prep for TEM from fixation through embedding. This general protocol applies to other specimens that require embedding for cross-sectioning. Photos show the mPrep capsule reagent exchange process and how mPrep TEM blocks are faced and sectioned while still in capsules. Supplies and the reagent formulae are listed.

This application note presents a fast, fully-automated method of tissue processing. Mammalian kidney was prepared for TEM with the mPrep ASP-1000 programmed for highly efficient rapid-agitation. Reagent processing from postfix rinse through 100% resin infiltration was accomplished in just 45 minutes.

Required

mPrep/s capsules to hold specimens

An insertion tool is required to insert, remove and adjust the mPrep/s screen. There are two options:

  •  mPrep/s Insertion tool
  •  mPrep/s Workstation (w/integrated insertion tool): required to orient specimens using the pinch method. Recommended because of its additional benefits
    • Precise specimen orientation in seconds
    • Wet samples easy to handle
    • Single focal plane keeps specimens in focus
    • Simultaneous loading of capsules onto pipettor for reagent processing

mPrep/f couplers to prevent introducing damaging reagents into pipettors

Pipettor (such as a Gilson P200 volume) to enable reagent pipetting into mPrep/s capsules

mPrep/bench to seal capsule bottoms during long reagent treatments and resin curing

Recommended

Reagent reservoirs

Microwell plates

Gilson Pipetman Neo (P200) pipettors bundled with mPrep/s capsules

mPrep/s™ Pipettor Kit single-channel

  • Single-channel pipettor
  • mPrep/s capsule pack
  • 96 pipette tips

mPrep/s™ Pipettor Kit eight-channel

  • Eight-channel pipettor
  • mPrep/s capsule pack
  • 96 pipette tips

Publications

Goodman S. and Kostrna M (2011). Reducing Reagent Consumption and Improving Efficiency of Specimen Fixation and Embedding, Grid Staining and Archiving using mPrep™ Capsule Processing. Microscopy and Microanalysis, 17 (Suppl. 2) , pp 1174-1175. doi:10.1017/S143192761100674X.

This short proceedings paper introduced the mPrep System at Microscopy and Microanalysis 2011. The paper provides an overview of the mPrep System for TEM specimen preparation and TEM grid staining. The System is compared to conventional processing for laboratory efficiency and reagent consumption.

McClain M. (2014) High Throughput Multi Parameter TEM Chemical Processing Protocol Development with the mPrep-s Capsule System: Schmidtea mediterranea. Microsc. Microanal. 20 (Suppl 3), 2014: 1288-9.   doi:10.1017/S1431927614008174

McClain simultaneously prepared 16 tissue specimens for TEM using 16 protocols to establish an optimal fixation and embedding protocol. This study varied buffer molarity, en bloc stains, and embedding resins. Reagent processing was conducted simultaneously using multi-channel pipettes and microtiter plates. The author reported, “The mPrep System facilitated the controlled and consistent testing of many conditions, which greatly increased the chances of obtaining optimized specimen preservation in one processing run.”

McClain M. (2016) Bringing Speed and Automation Together with the ASP1000. Microsc. Microanal. 2016. Post deadline poster #49.

McClain developed a TEM tissue fixation and embedding protocol that utilized high speed agitation on the ASP-1000 to prepare planarian worm specimens for resin curing that required only 3 hours. The results were comparable to a 7-day embedding protocol that was previously used to prepare these worm specimens.

Goodman SL, Wendt KD, Kostrna MS, Radi C (2015) Capsule-Based Processing and Handling of Electron Microscopy Specimens and Grids. Microscopy Today, 23(5): 30-7. doi:10.1017/S155192951500076

Authors from three institutions prepared animal and plant tissues for TEM. They used mPrep/s capsules for fixation through embedding, and mPrep/g capsules for grid staining. Researches assessed reagent consumption and labor efficiency of mPrep processing compared to traditional processing (using vials, embedding molds and droplet staining). The findings included: mPrep/s specimen processing reduced reagent consumption ten-fold and the number of hands-on operations seven-fold. mPrep/g grid staining reduced hands-on operations six-fold.


Videos

Preparing Embedded TEM Specimens for Sectioning

This video tutorial shows how to trim and face an embedded block inside an mPrep/s capsule. The capsule is clamped into a microtome chuck for trimming, facing and sectioning.

1 minutes 17 seconds

Using the mPrep Workstation for Oriented and Unoriented Specimens

The tutorial starts by introducing the mPrep Workstation. Then it demonstrates how to load and orient specimens into mPrep capsules. Three methods are shown: pinch, compression and cup.

5 minutes 19 seconds


Instructions

A single page graphical display of the three steps to insert specimens into mPrep/s capsules using the mPrep/s insertion tool. Use this guide as a lab reference poster.

A single page graphical display of the three steps to orient and load specimens into mPrep/s capsules. Both compression and pinch methods are shown. Use this guide as a lab reference poster.

Instructions for using mPrep/s capsules for specimen handling and processing.

Print this 96-well plate template to organize protocols into columns and rows with a microtiter 96-well plate.

This technical note introduces using mPrep capsule labeling for tracking specimens and grids with a data management system. Topics addressed include mPrep labels, 2D datamatrix barcodes, barcode readers and laboratory information management systems.


mPrep specimen products

mPrep/s™ Capsule Pack

8 capsules, 12 screens, storage box, labels

Product SKU: 22200

$48.00

mPrep/s™ Capsules – 96 count

96 capsules, screens & blank label sets

Product SKU: 22500

$576.00

mPrep/s™ Insertion Tool

Inserts, removes and adjusts screens in mPrep/s capsules

Product SKU: 32010

$25.00

mPrep/s™ Workstation

Prepare specimens and insert into mPrep/s capsules with or without orientation using integrated screen insertion tool

Product SKU: 42100

$950.00

mPrep/Bench™ model 96s

96-well silicone rack for mPrep capsules

Product SKU: 34000

$45.00

mPrep/f 30 Standard Filter Coupler™

16 couplers

Product SKU: 31500

$48.00

mPrep/f 13 Extreme Filter Coupler™

16 couplers

Product SKU: 31600

$48.00

mPrep™ CPD Holder

Holds up to 6 mPrep/s or mPrep/g capsules in Tousimis® CPD apparatus

Product SKU: 33010

$225.00

ASP-1000 mPrep Automatic Specimen Processor

Programable specimen and grid preparation.     Request a Quotation

Product SKU: ASP1000

Reagent Reservoirs

Chemically resistant 15ml reservoirs, 50 per pack

Product SKU: 52001

$22.00

Microwell Plates

Chemically resistant 96-well plates, 10 per pack

Product SKU: 51001

$39.00

mPrep/s™ Pipettor Kit eight-channel

eight-channel pipettor, mPrep/s capsule pack, 96 pipette tips

Product SKU: 43250

$686.00

mPrep/s™ Pipettor Kit single-channel

single-channel pipettor, mPrep/s capsule pack, 96 pipette tips

Product SKU: 43150

$351.00

Staining tissue sections on TEM grids

The mPrep System streamlines the entire tissue preparation and grid staining workflow.

Problems

Staining tissue sections on grids requires using fine forceps to transfer grids into grid boxes, onto stain droplets or staining apparatuses, back into grid boxes, and eventually onto the TEM stage. As a result, microscopists experience multiple challenges:

  • Grids are easily damaged
  • Handling grids is tedious
  • Grids are easily lost or mixed up
  • Difficulty achieving uniform staining times across multiple grids
  • Stains exposed to air can be contaminated, precipitate, and evaporate

 

mPrep solutions

  • Encapsulated grids reduce handling and damage.
  • Grids may only be directly handled at the microtome and the microscope
  • Grids in mPrep/g capsules are always labeled
  • Simultaneous processing assures uniform timing
  • Enclosed capsules reduce and can even eliminate air exposure

Learn more about staining tissue sections on TEM grids in the application notes below.

Heart (TEM)

Brain - Vampire bat (TEM)

Application Notes

This application note illustrates staining tissue sections on TEM grids using mPrep/g capsules with pipettors and common lab supplies. The protocol shows uranyl acetate and lead citrate staining of kidney and brain sections, although the methods apply to other grid staining procedures. Required supplies and the reagent formulae are listed.

Required

mPrep/g capsules to hold TEM grids

mPrep/f couplers to prevent introducing damaging reagents into pipettors

Pipettor (such as a Gilson P200 volume) to enable reagent pipetting into mPrep/s capsules

Recommended

Reagent reservoirs

Microwell plates

mPrep/Bench to seal capsule bottoms during long reagent treatments

Gilson Pipetman Neo (P200) pipettors bundled with mPrep/g capsules

mPrep/g Pipettor Kit single-channel

  • Single-channel pipettor
  • mPrep/s capsule pack
  • 96 pipette tips

mPrep/g Pipettor Kit eight-channel

  • Eight-channel pipettor
  • mPrep/g capsule pack
  • 96 pipette tips

Publications

Goodman SL, Wendt KD, Kostrna MS, Radi C (2015) Capsule-Based Processing and Handling of Electron Microscopy Specimens and Grids. Microscopy Today, 23(5): 30-7. doi:10.1017/S155192951500076

Authors from three institutions prepared animal and plant tissues for TEM. They used mPrep/s capsules for fixation through embedding, and mPrep/g capsules for grid staining. Researches assessed reagent consumption and labor efficiency of mPrep processing compared to traditional processing (using vials, embedding molds and droplet staining). The findings included: mPrep/s specimen processing reduced reagent consumption ten-fold and the number of hands-on operations seven-fold. mPrep/g grid staining reduced hands-on operations six-fold.

Radi C. (2015) Improving the Reliability, Ease, and Efficiency of Section Staining in a Diagnostic Laboratory with the mPrep/g System for TEM Grid Processing. Microsc. Microanal. 21 (Suppl 3), 2015: 725-6. doi: 10.1017/S1431927615004420

Radi discusses the necessity for efficiently preparing and documenting TEM grids in a clinical diagnostic laboratory setting. He then compares the droplet method to the mPrep system process for uranyl acetate and lead citrate staining of tissue sections . He reports that, “In addition to providing more consistent results, mPrep/g processing was also easier and more efficient.” And notes that “ . . . the time to prepare, store, document and stain each grid took only seconds compared to minutes using the droplet method.”

Strader TE, Goodman SL. (2015) Automated, Programmable Processing of Specimens and Grids with the mPrep ASP-1000. Microsc. Microanal. 21 (Suppl 3), 2015: 11-2. doi: 10.1017/S1431927615000859

Strader and Goodman introduce the mPrep Automated Specimen Processor (ASP), used for simultaneous sample preparation using mPrep capsules. The ASP-1000 prepares TEM and SEM specimens, and stains and labels TEM grids. The paper discusses the ASP-1000 features, including flexibility to efficiently perform routine and complicated tasks. The paper includes pictures of the instrument and examples of heart and skin tissue specimens prepared using mPrep capsule processing.

Benmeradi N., Payre B., Goodman S.L. (2015) Easier and Safer Biological Staining: High Contrast UranyLess Staining of TEM Grids using mPrep/g Capsules. Microsc. Microanal. 21 (Suppl 3), 2015: 721-2. doi: 10.1017/S1431927615004407

This paper demonstrates how a new lanthanide-based heavy metal stain, when used with mPrep/g capsules, provides high contrast tissue section and negative staining with results that are comparable or possibly superior to conventional uranyl acetate staining. Examples show TEM thin section staining of 5 animal tissues and one plant tissue, and a negative stained TEM bacteriophage specimen.

Goodman S. and Kostrna M (2011). Reducing Reagent Consumption and Improving Efficiency of Specimen Fixation and Embedding, Grid Staining and Archiving using mPrep™ Capsule Processing. Microscopy and Microanalysis, 17 (Suppl. 2) , pp 1174-1175. doi:10.1017/S143192761100674X.

This short proceedings paper introduced the mPrep System at Microscopy and Microanalysis 2011. The paper provides an overview of the mPrep System for TEM specimen preparation and TEM grid staining. The System is compared to conventional processing for laboratory efficiency and reagent consumption.


Videos

Specimen and Grid Reagent Processing for TEM and SEM

This video tutorial demonstrates how to use mPrep capsules with multi-channel pipettors and common lab ware to simultaneously deliver reagents to multiple specimens and grids. The video demonstrates TEM fixation and embedding. Reagent processing procedures are similar for all specimen types and for grids with mPrep capsules .

6 minutes 4 seconds


Instructions

Instructions for using mPrep/g capsules for staining sections on grids with uranyl acetate and lead citrate.

Instructions for using mPrep/g capsules for grid handling and processing.

This technical note introduces using mPrep capsule labeling for tracking specimens and grids with a data management system. Topics addressed include mPrep labels, 2D datamatrix barcodes, barcode readers and laboratory information management systems.

A graphical display illustrating how to load grids into mPrep/g capsules, and how to use pipettors to stain or label grids. Use this guide as a lab reference poster.


mPrep grid products

mPrep/Bench™ model 96s

96-well silicone rack for mPrep capsules

Product SKU: 34000

$45.00

mPrep/g™ Capsule Pack

16 capsules, storage box, labels

Product SKU: 21300

$48.00

mPrep/g™ Capsules – 96 count

96 capsules, 96 label sets

Product SKU: 21500

$288.00

mPrep/f 30 Standard Filter Coupler™

16 couplers

Product SKU: 31500

$48.00

mPrep/f 13 Extreme Filter Coupler™

16 couplers

Product SKU: 31600

$48.00

mPrep™ CPD Holder

Holds up to 6 mPrep/s or mPrep/g capsules in Tousimis® CPD apparatus

Product SKU: 33010

$225.00

ASP-1000 mPrep Automatic Specimen Processor

Programable specimen and grid preparation.     Request a Quotation

Product SKU: ASP1000

Reagent Reservoirs

Chemically resistant 15ml reservoirs, 50 per pack

Product SKU: 52001

$22.00

Microwell Plates

Chemically resistant 96-well plates, 10 per pack

Product SKU: 51001

$39.00

mPrep/g™ Pipettor Kit eight-channel

eight-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43200

$686.00

mPrep/g™ Pipettor Kit single-channel

single-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43100

$351.00

Negative staining of macromolecules

The mPrep system simplifies the preparation of macromolecular and particulate specimens for TEM.

Problems

Negative stain specimen preparation requires fine forceps to transfer filmed grids across droplets of macromolecule or particle suspensions, negative stains, and rinses.  This presents microscopists with multiple challenges:

  • Grids and support films are easily damaged
  • Handling grids is tedious
  • Grids are easily lost or mixed up
  • Limited quantities of specimen solutions may be available
  • Specimen and reagent solutions may evaporate, precipitate or be contaminated
  • Inconsistent adsorption and staining times across multiple grids

 

 mPrep  solutions

  • Encapsulated grids reduce handling and  damage
  • Grids with specimens may require direct handling only at the microscope
  • Grids in mPrep/g capsules are always labeled
  • Capsules require only 35 μl to prepare two grids
  • Enclosed capsules reduce or eliminate air exposure
  • Simultaneous processing assures uniform timing

DNA RecA (TEM negative stain)

Application Notes

This application note illustrates the preparation of macromolecular specimens for negative stain TEM using mPrep/g capsules, pipettors and Formvar®-filmed TEM grids. The example is a DNA-RecA complex, although the protocol is applicable to many other specimens. The specimen preparation and microscopy was performed in the Department of Biochemistry at the University of Wisconsin – Madison by Mr. Jay Campbell and Dr. Sindhu Chitteni-Pattu, in the laboratory of Dr. Michael M Cox.

This application note illustrates staining tissue sections on TEM grids using mPrep/g capsules with pipettors and common lab supplies. The protocol shows uranyl acetate and lead citrate staining of kidney and brain sections, although the methods apply to other grid staining procedures. Required supplies and the reagent formulae are listed.

Required

mPrep/g capsules to hold TEM grids

mPrep/f couplers to prevent introducing damaging reagents into pipettors

Pipettor (such as a Gilson P200 volume) to enable reagent pipetting into mPrep/s capsules

Recommended

Reagent reservoirs
Microwell plates
mPrep/bench to seal capsule bottoms during long reagent treatments

Gilson Pipetman Neo (P200) pipettors bundled with mPrep/g capsules.

mPrep/g Pipettor Kit single-channel

  • Single-channel pipettor
  • mPrep/s capsule pack
  • 96 pipette tips

mPrep/g Pipettor Kit eight-channel

  • Eight-channel pipettor
  • mPrep/g capsule pack
  • 96 pipette tips

Publications

Benmeradi N., Payre B., Goodman S.L. (2015) Easier and Safer Biological Staining: High Contrast UranyLess Staining of TEM Grids using mPrep/g Capsules. Microsc. Microanal. 21 (Suppl 3), 2015: 721-2. doi: 10.1017/S1431927615004407

This paper demonstrates how a new lanthanide-based heavy metal stain, when used with mPrep/g capsules, provides high contrast tissue section and negative staining with results that are comparable or possibly superior to conventional uranyl acetate staining. Examples show TEM thin section staining of 5 animal tissues and one plant tissue, and a negative stained TEM bacteriophage specimen.


Videos

Specimen and Grid Reagent Processing for TEM and SEM

This video tutorial demonstrates how to use mPrep capsules with multi-channel pipettors and common lab ware to simultaneously deliver reagents to multiple specimens and grids. The video demonstrates TEM fixation and embedding. Reagent processing procedures are similar for all specimen types and for grids with mPrep capsules .

6 minutes 4 seconds


Instructions

Instructions for using mPrep/g capsules for grid handling and processing.

Print this 96-well plate template to organize protocols into columns and rows with a microtiter 96-well plate.

A graphical display illustrating how to load grids into mPrep/g capsules, and how to use pipettors to stain or label grids. Use this guide as a lab reference poster.


mPrep grid products

mPrep/Bench™ model 96s

96-well silicone rack for mPrep capsules

Product SKU: 34000

$45.00

mPrep/g™ Capsule Pack

16 capsules, storage box, labels

Product SKU: 21300

$48.00

mPrep/g™ Capsules – 96 count

96 capsules, 96 label sets

Product SKU: 21500

$288.00

mPrep/f 30 Standard Filter Coupler™

16 couplers

Product SKU: 31500

$48.00

mPrep/f 13 Extreme Filter Coupler™

16 couplers

Product SKU: 31600

$48.00

mPrep™ CPD Holder

Holds up to 6 mPrep/s or mPrep/g capsules in Tousimis® CPD apparatus

Product SKU: 33010

$225.00

ASP-1000 mPrep Automatic Specimen Processor

Programable specimen and grid preparation.     Request a Quotation

Product SKU: ASP1000

Reagent Reservoirs

Chemically resistant 15ml reservoirs, 50 per pack

Product SKU: 52001

$22.00

Microwell Plates

Chemically resistant 96-well plates, 10 per pack

Product SKU: 51001

$39.00

mPrep/g™ Pipettor Kit eight-channel

eight-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43200

$686.00

mPrep/g™ Pipettor Kit single-channel

single-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43100

$351.00

Preparing viruses on TEM grids

The mPrep system facilitates preparation of viruses and other microorganisms for TEM examination, including  those prepared in Biological Safety Level 3 or 4 laboratory suites.

Problems

Virus sample preparation for TEM requires using fine forceps to transfer filmed grids throughout the process.  Grids boxes may require decontamination prior to transfer to the TEM.

  • Grids and support films are easily damaged
  • Grids are easily lost or mixed up
  • Sharp forceps are a puncture risk to protective clothing
  • Grid handling is especially difficult wearing BSL3/4 positive pressure suits
  • Limited quantities of specimen solutions may be available
  • Specimen and reagent solutions exposed to air may evaporate, precipitate or be contaminated
  • Grids and grid storage containers require decontamination
  • Adsorption and staining times may be inconsistent across multiple grids

 

mPrep solutions

 

  • Encapsulated grids reduce handling and damage
  • Grids in mPrep/g capsules are always labeled
  • Forceps are not used for virus preparation
  • Direct handling of grids may require handling only at the microscope
  • Capsules require only 35 μl to prepare two grids
  • Enclosed capsules reduce or eliminate air exposure
  • Grids in capsules are decontaminated as a unit
  • Simultaneous processing assures uniform timing

T6 Phage (negative stain)

Ebola virus (negative stain)

Application Notes

This application note shows the negative stain TEM preparation of pathogenic Ebola virus in high containment BSL4 labs. Scientists used mPrep/g capsules to eliminate grid handling in the BSL suite. This resulted in easier, safer processing and more reproducible sample preparation.  Specimen preparation and microscopy were performed by the US Army Medical Research Institute of Infectious Diseases (USAMRIID) Pathology Division, at Ft. Detrick, MD.

Required

mPrep/g capsules to hold TEM grids
mPrep/f 13 extreme couplers to prevent introducing damaging reagents or viruses into pipettors
Pipettor (such as a Gilson P200 volume) to enable reagent pipetting into mPrep/s capsules

Recommended

Reagent reservoirs

Microwell plates

mPrep/bench to seal capsule bottoms during long reagent treatments

Gilson Pipetman Neo (P200) pipettors bundled with mPrep/g capsules.

mPrep/g Pipettor Kit single-channel

  • Single-channel pipettor
  • mPrep/s capsule pack
  • 96 pipette tips

mPrep/g Pipettor Kit eight-channel

  • Eight-channel pipettor
  • mPrep/g capsule pack
  • 96 pipette tips

Publications

Benmeradi N., Payre B., Goodman S.L. (2015) Easier and Safer Biological Staining: High Contrast UranyLess Staining of TEM Grids using mPrep/g Capsules. Microsc. Microanal. 21 (Suppl 3), 2015: 721-2. doi: 10.1017/S1431927615004407

This paper demonstrates how a new lanthanide-based heavy metal stain, when used with mPrep/g capsules, provides high contrast tissue section and negative staining with results that are comparable or possibly superior to conventional uranyl acetate staining. Examples show TEM thin section staining of 5 animal tissues and one plant tissue, and a negative stained TEM bacteriophage specimen.

Monninger MK, Nguessan CA, Blancett CD, Kuehl KA, Rossi CA, Olschner SP, Williams PL, Goodman SL, Sun MG.  (2016)  Improved Virus Specimen Preparation for Transmission Electron Microscopy using mPrep/g Capsules: Applications in BSL3-4 Laboratories.  Ft. Detrick Science Fair, 5 May 2016.

This poster shows how mPrep/g capsules enable the easy preparation of pathogenic viruses in BSL4 biocontainment. The authors from USAMRIID (US Army Medical Research Institute of Infectious Diseases) prepared Zaire Ebola virus and Chikungunya virus with glutaraldehyde inactivation and negative staining. Samples were also prepared of Ebola nano virus like particles and murine Leukemia virus like particles. Morphology of the viruses and virus like particles were assessed with and without osmium tetroxide vapor inactivation, and with uranyl acetate and phosphotungstic acid negative staining.

Monninger MK, Nguessan CA, Blancett CD, Goodman SL, Sun MG. (2016) Improved Virus Specimen Preparation for Transmission Electron Microscopy using mPrep/g Capsules: Applications in BSL3-4 Laboratories.  Microsc. Microanal. 22 (Suppl 3), 2016: 1164. doi:10.1017/S1431927616006668

This paper shows how mPrep/g capsules enables easy and reproducible preparation of highly pathogenic viruses for TEM by eliminating grid handling in BSL3-4 biocontainment suites. Examples are shown of negative stained Zaire Ebola virus and murine leukemia virus-like particles. A pictorial diagrams the preparation protocol inside and outside the biocontainment suite. The authors report that the reduced effort, consistent quality, and simultaneous preparation of multiple grids makes this method equally useful for labs working with non-pathogenic viruses.

Blancett CD, Fetterer DP, Koistinen KA, Morazzani EM, Monninger MK, Piper AE, Kuehl KA,  Glass PJ, Sun MG. (2016) Using Scanning Transmission Electron Microscopy (STEM) for Accurate Virus Dosing Quantification. Microsc. Microanal. 22 (Suppl 3), 2016: 1162. doi:10.1017/S1431927616006656

Blancett and co-authors use mPrep/g grid capsules to reproducibly and accurately quantify unknown virus concentrations. Virus suspensions are mixed with a known concentration of nano-gold particles and applied to Formvar-carbon grids using mPrep/g capsules and pipettors. The grids are then imaged with scanning transmission electron microscopy (STEM) followed by automatic image analysis to quantify viral concentration by counting the ratio of viruses to nano-gold particles. They report that mPrep/g preparation improves enumeration accuracy, and that assay results are consistent with virus plaque assays.

Monninger MK, Nguessan CA, Blancett CD, Kuehl KA, Rossi CA, Olschner SP, Williams PL,  Goodman SL,  Sun MG. (2016) Preparation of viral samples within biocontainment for ultrastructural analysis: Utilization of an innovative processing capsule for negative staining. Journal of Virological Methods 238 (2016) 70–76. doi.org/10.1016/j.jviromet.2016.10.005

This study demonstrates how to prepare Ebola virus, chikungunya virus, murine leukemia virus and virus-like particle in BSL3-4 biocontainment suites using mPrep/g capsules. The study also compares virus inactivation using glutaraldehyde fixation prior to grid preparation and using osmium tetroxide vapor treatment after grid preparation. In addition, uranyl acetate and phosphotungstatic acid negative staining are compared. The authors conclude that mPrep/g capsule preparation saves time and effort and produces more consistent high-quality TEM images than manual droplet grid preparation. The study also concludes that uranyl acetate and phosphotungstatic acid staining provide comparable results, and that osmium tetroxide vapor decontamination is faster than glutaraldehyde decontamination but can reduce image quality.



Instructions

Instructions for using mPrep/g capsules for grid handling and processing.

Print this 96-well plate template to organize protocols into columns and rows with a microtiter 96-well plate.

A graphical display illustrating how to load grids into mPrep/g capsules, and how to use pipettors to stain or label grids. Use this guide as a lab reference poster.

A graphical display that illustrates how to easily prepare virus specimens for negative stain TEM in Biological Safety Level 3-4, and other laboratories. Use this guide as a lab reference poster.


mPrep grid products

mPrep/Bench™ model 96s

96-well silicone rack for mPrep capsules

Product SKU: 34000

$45.00

mPrep/g™ Capsule Pack

16 capsules, storage box, labels

Product SKU: 21300

$48.00

mPrep/g™ Capsules – 96 count

96 capsules, 96 label sets

Product SKU: 21500

$288.00

mPrep/f 30 Standard Filter Coupler™

16 couplers

Product SKU: 31500

$48.00

mPrep/f 13 Extreme Filter Coupler™

16 couplers

Product SKU: 31600

$48.00

mPrep™ CPD Holder

Holds up to 6 mPrep/s or mPrep/g capsules in Tousimis® CPD apparatus

Product SKU: 33010

$225.00

ASP-1000 mPrep Automatic Specimen Processor

Programable specimen and grid preparation.     Request a Quotation

Product SKU: ASP1000

Reagent Reservoirs

Chemically resistant 15ml reservoirs, 50 per pack

Product SKU: 52001

$22.00

Microwell Plates

Chemically resistant 96-well plates, 10 per pack

Product SKU: 51001

$39.00

mPrep/g™ Pipettor Kit eight-channel

eight-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43200

$686.00

mPrep/g™ Pipettor Kit single-channel

single-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43100

$351.00

Immunogold labeling (IGL)

Immunogold labeling (IGL) or immunolabeling is a powerful methodology for localizing structures in biological specimens.

Problems

Immunolabeling tissue sections on TEM grids using “droplet” methods requires the timed transfer of multiple grids through a dozen reagents in a process that can take hours or even a full day. As a result, microscopists experience multiple challenges:

  • Difficultly achieving consistent reagent timing, which limits experimental reproducibility
  • Grids are easily damaged, lost, or mixed up
  • IGL reagents exposed to air can evaporate, precipitate or become contaminated
  • IGL reagents and antibodies are expensive

 

mPrep solutions

  • The mPrep System addresses these challenges:
    • Simultaneous processing assures identical reagent timing and reproducible results
    • Encapsulated grids are identified and protected from loss or damage
    • Enclosed capsules limit air exposure
    • mPrep/g capsules require only 20 ul per grid

 

Automated IGL with the mPrep ASP-1000

With the ASP-1000 the entire immunolabeling process is automated. Simply set up the reagents, start the program, and walk away for the rest of the day or overnight. Use with mPrep/g capsules for grids, and mPrep/s capsules for tissue en bloc IGL.

 

Manual IGL with a multi-channel pipettor

Using a multichannel pipettor ensures that reagent incubation timing is identical for all grids. Because grids are safely encapsulated reagent exchanges are fast and easy. Use with mPrep/s capsules for tissue en bloc IGL.

6 nm IGL, endosome complex, C. elegans

6 nm IGL, endosome complex, C. elegans

Application Notes

This brief technical note illustrates automation of immunogold labeling using the mPrep ASP-1000 automated specimen processor. Researchers at the University of Wisconsin prepared C. elegans tissue sections on TEM grids and then used the ASP-1000 to immunolabel an endosomal sorting component with 6 nm gold. The entire 7.5 hour immunolabeling protocol was automatically accomplished.  This note also illustrates automated application of 10 nm gold fiducial markers onto thick sections for tomography.

Required

mPrep/g capsules to hold TEM grids
mPrep/f couplers to prevent introducing reagents into the ASP-1000 or pipettors

ASP-1000 for automated immunogold labeling

Pipettor (such as a Gilson P200 volume) to enable reagent pipetting into mPrep/s capsules

mPrep/s capsules to hold tissue samples for en bloc IGL

Recommended

Microwell plates to hold reagents

Foil seals to cover reagents until use

Example: 1.5 mil thick  pierceable AlumaSeal II Film for PCR plates (Contact us for more information)

Publications

Frankel EB August BK, Audhya A, Strader TE (2016) Fully-automated Immunogold Labeling of Resin Embedded Specimens and On-grid Deposition of Gold Fiducial Particles. Microsc. Microanal, 22 (Suppl 3), 1010-1. doi 10.1017/S1431927616005894

This proceedings paper and presented poster demonstrates fully automated immunogold labeling of an endosomal sorting complex in a C. elegans 1-cell stage embryo. The authors report that immunogold labeling using the mPrep ASP-1000 was highly reproducible across numerous samples. This work also demonstrates automated application of gold colloids to thick sections on grids to provide fiducial markers for electron tomography. The authors report that gold fiducial particles were adsorbed evenly over the grid surfaces allowing for highly accurate registration of tilt-series images.



Instructions

Instructions for using mPrep/g capsules for grid handling and processing.

A graphical display illustrating how to load grids into mPrep/g capsules, and how to use pipettors to stain or label grids. Use this guide as a lab reference poster.

Protocol for immuno-labeling grids with mPrep/g


mPrep grid products

mPrep/Bench™ model 96s

96-well silicone rack for mPrep capsules

Product SKU: 34000

$45.00

mPrep/g™ Capsule Pack

16 capsules, storage box, labels

Product SKU: 21300

$48.00

mPrep/g™ Capsules – 96 count

96 capsules, 96 label sets

Product SKU: 21500

$288.00

mPrep/f 30 Standard Filter Coupler™

16 couplers

Product SKU: 31500

$48.00

mPrep/f 13 Extreme Filter Coupler™

16 couplers

Product SKU: 31600

$48.00

mPrep™ CPD Holder

Holds up to 6 mPrep/s or mPrep/g capsules in Tousimis® CPD apparatus

Product SKU: 33010

$225.00

ASP-1000 mPrep Automatic Specimen Processor

Programable specimen and grid preparation.     Request a Quotation

Product SKU: ASP1000

Reagent Reservoirs

Chemically resistant 15ml reservoirs, 50 per pack

Product SKU: 52001

$22.00

Microwell Plates

Chemically resistant 96-well plates, 10 per pack

Product SKU: 51001

$39.00

mPrep/g™ Pipettor Kit eight-channel

eight-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43200

$686.00

mPrep/g™ Pipettor Kit single-channel

single-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43100

$351.00

Colloidal gold fiducial markers for 3-D Tomography

Accurate image alignment is essential for 3-D image reconstruction in TEM tomography. Colloidal gold particles are used as fiducial markers to align the different tilt angle images of thick sections.

Problems

For successful image reconstruction colloidal gold fiducial markers must be distributed uniformly over both sides of the grid. The common protocol is to apply a droplet of a colloidal gold suspension to each grid, and then wick off the droplet after a short adsorption time. The result is that microscopists experience multiple challenges:

  • Inconsistent results from grid to grid
  • Uneven particle distributions on each grid and each grid side
  • Time consuming to prepare multiple grids
  • Grids are easily damaged, lost, or mixed up

 

mPrep solutions

The mPrep System addresses these challenges:

  • Consistent results on every grid
  • Uniform particle distribution on both grid sides
  • Automated or manual simultaneous preparation of up to 16 grids
  • Encapsulated grids are protected from damage or loss, and are always identified

 

Automated with the mPrep ASP-1000

Application of colloidal gold fiducial particles onto grids is fully automated with the mPrep ASP-1000. Colloidal gold particles are applied simultaneously to both sides of up to 16 grids. Automated agitation ensures consistent results.

 

Manual IGL with a multi-channel pipettor

Using mPrep/g capsules with an 8-channel pipettor, colloidal gold fiducial particles are applied simultaneously to both sides of up to 16 grids. With a multichannel pipettor, manual agitation ensures consistent results for multiple grids prepared simultaneously.

10 nm Au fiducial particle, 250 nm section

10 nm Au fiducial particle, 250 nm section

Required

mPrep/g capsules to hold TEM grids

mPrep/f couplers to prevent introducing reagents into the ASP-1000 or pipettors

ASP-1000 for automated immunogold labeling

Pipettor (such as a Gilson P200 volume) to enable reagent pipetting into mPrep/s capsules

Recommended

Microwell plates to hold reagents

Publications

Frankel EB August BK, Audhya A, Strader TE (2016) Fully-automated Immunogold Labeling of Resin Embedded Specimens and On-grid Deposition of Gold Fiducial Particles. Microsc. Microanal, 22 (Suppl 3), 1010-1. doi 10.1017/S1431927616005894

This proceedings paper and presented poster demonstrates fully automated immunogold labeling of an endosomal sorting complex in a C. elegans 1-cell stage embryo. The authors report that immunogold labeling using the mPrep ASP-1000 was highly reproducible across numerous samples. This work also demonstrates automated application of gold colloids to thick sections on grids to provide fiducial markers for electron tomography. The authors report that gold fiducial particles were adsorbed evenly over the grid surfaces allowing for highly accurate registration of tilt-series images.



Instructions

Instructions for using mPrep/g capsules for grid handling and processing.

A graphical display illustrating how to load grids into mPrep/g capsules, and how to use pipettors to stain or label grids. Use this guide as a lab reference poster.

Protocol for immuno-labeling grids with mPrep/g


mPrep grid products

mPrep/Bench™ model 96s

96-well silicone rack for mPrep capsules

Product SKU: 34000

$45.00

mPrep/g™ Capsule Pack

16 capsules, storage box, labels

Product SKU: 21300

$48.00

mPrep/g™ Capsules – 96 count

96 capsules, 96 label sets

Product SKU: 21500

$288.00

mPrep/f 30 Standard Filter Coupler™

16 couplers

Product SKU: 31500

$48.00

mPrep/f 13 Extreme Filter Coupler™

16 couplers

Product SKU: 31600

$48.00

mPrep™ CPD Holder

Holds up to 6 mPrep/s or mPrep/g capsules in Tousimis® CPD apparatus

Product SKU: 33010

$225.00

ASP-1000 mPrep Automatic Specimen Processor

Programable specimen and grid preparation.     Request a Quotation

Product SKU: ASP1000

Reagent Reservoirs

Chemically resistant 15ml reservoirs, 50 per pack

Product SKU: 52001

$22.00

Microwell Plates

Chemically resistant 96-well plates, 10 per pack

Product SKU: 51001

$39.00

mPrep/g™ Pipettor Kit eight-channel

eight-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43200

$686.00

mPrep/g™ Pipettor Kit single-channel

single-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43100

$351.00

Preparing nanoparticles on TEM grids

The mPrep system simplifies the preparation of nanoparticles for TEM.

Problems

Nanoparticle specimen prep requires  forceps to transfer filmed grids through multiple reagents. Grids are stored in grid boxes until TEM imaging.

  • Grids and support films are easily damaged
  • Grids are easily lost or mixed up
  • Limited quantities of specimen solutions may be available
  • Specimen and reagent solutions may evaporate, precipitate or be contaminated
  • Adsorption and staining times may be inconsistent across multiple grids

 

mPrep solutions

  • Encapsulated grids reduce handling and damage
  • Grids in mPrep/g capsules are always labeled
  • Capsules require only 35 μl to prepare two grids
  • Enclosed capsules reduce or eliminate air exposure
  • Simultaneous processing assures uniform timing

Nanoparticles (TEM negative stain)

Application Notes

This illustrated protocol shows the preparation of nanoparticle specimens for negative stain TEM using Formvar®-filmed grids in mPrep/g capsules. Eight different formulations of ~100 nm drug delivery particles were prepared simultaneously for a pharmaceutical company to evaluate structure and uniformity. This general sample preparation method can be applied to other nano-scale specimens including nano-fibers and viruses.

This application note illustrates the preparation of macromolecular specimens for negative stain TEM using mPrep/g capsules, pipettors and Formvar®-filmed TEM grids. The example is a DNA-RecA complex, although the protocol is applicable to many other specimens. The specimen preparation and microscopy was performed in the Department of Biochemistry at the University of Wisconsin – Madison by Mr. Jay Campbell and Dr. Sindhu Chitteni-Pattu, in the laboratory of Dr. Michael M Cox.

Required

mPrep/g capsules to hold TEM grids
mPrep/f couplers to prevent introducing damaging reagents into pipettors
Pipettor (such as a Gilson P200 volume) to enable reagent pipetting into mPrep/s capsules

Recommended

Reagent reservoirs

Microwell plates

mPrep/bench to seal capsule bottoms during long reagent treatments

Gilson Pipetman Neo (P200) pipettors bundled with mPrep/g capsules

mPrep/g Pipettor Kit single-channel

  • Single-channel pipettor
  • mPrep/s capsule pack
  • 96 pipette tips

mPrep/g Pipettor Kit eight-channel

  • Eight-channel pipettor
  • mPrep/g capsule pack
  • 96 pipette tips

Product Descriptions

Publications

Vierrether O, TerBush J, Wisner C (2016) Nano-Particle TEM Sample Preparation Primer. Microsc. Microanal. 22 (Suppl 3), 2016: 1914. doi:10.1017/S1431927616010412

This paper and poster provides a guide to selecting grid and support film types for preparing nanoparticles for TEM, and also compares droplet preparation with mPrep ASP-1000 automated processing. The authors concluded that lacey silicon monoxide/Formvar films on 300 mesh grids proved to the be the best for the nanoparticles examined. They also concluded that mPrep automation reduced sample agglomeration thereby allowing for single particle imaging, and this could also be achieved with mPrep/g capsules and manual pipetting.

Goodman S.L. (2015) Efficient and Documented Preparation of Pharmaceutical Particles for Correlative Microscopy Analyses using mPrep Capsule Processing Microsc. Microanal. 21 (Suppl 3), 2015: 393-4. doi: 10.1017/S1431927615002767

Using mPrep capsules, Goodman prepared two types of pharmaceutical particles for characterization. The mPrep capsules enabled simultaneous and GLP compliant sample preparation. Goodman used mPrep/g capsules to simultaneously prepare 16 grids for TEM from two formulations of siRNA drug delivery nanoparticles with four staining protocols. The grids were then analyzed for morphology and uniformity using TEM. In a separate study he simultaneously embedded four formulations of polymeric drug delivery micro-particles to create eight blocks for cross-sectioning. Thin sections were than analyzed for chemical composition using transmission FTIR and for elemental composition using EDS-SEM.

Jain R, Maric R (2014) Synthesis of nano-Pt onto ceria support as catalyst for water–gas shift reaction by Reactive Spray Deposition Technology. Applied Catalysis A: General, 475: 461–468.

Jain et al used mPrep/g capsules to prepare nanoparticle specimens for high resolution TEM characterization. They deposited reactive spray synthesized nano-Pt catalysts onto lacy carbon filmed 300 mesh TEM grids in mPrep/g capsules.

Jain R, Poyraz AS, Gamliel DP, Valla J, Suib SL, Maric R (2015) Comparative study for low temperature water-gas shift reaction on Pt/ceria catalysts: Role of different ceria supports. Applied Catalysis A: General, 507: 1–13.

Jain, et al used mPrep/g capsules to prepare nanoparticle specimens for high resolution TEM characterization. They tested and characterized different Pt on ceria catalyst using XRD, Raman, BET, SEM and TEM. For TEM imaging, the catalysts were deposited onto carbon filmed 300 mesh TEM grids using mPrep/g capsules.



Instructions

Instructions for using mPrep/g capsules for grid handling and processing.

Instructions for using mPrep/g capsules for staining sections on grids with uranyl acetate and lead citrate.

A graphical display illustrating how to load grids into mPrep/g capsules, and how to use pipettors to stain or label grids. Use this guide as a lab reference poster.


Product Descriptions

mPrep grid products

mPrep/Bench™ model 96s

96-well silicone rack for mPrep capsules

Product SKU: 34000

$45.00

mPrep/g™ Capsule Pack

16 capsules, storage box, labels

Product SKU: 21300

$48.00

mPrep/g™ Capsules – 96 count

96 capsules, 96 label sets

Product SKU: 21500

$288.00

mPrep/f 30 Standard Filter Coupler™

16 couplers

Product SKU: 31500

$48.00

mPrep/f 13 Extreme Filter Coupler™

16 couplers

Product SKU: 31600

$48.00

mPrep™ CPD Holder

Holds up to 6 mPrep/s or mPrep/g capsules in Tousimis® CPD apparatus

Product SKU: 33010

$225.00

ASP-1000 mPrep Automatic Specimen Processor

Programable specimen and grid preparation.     Request a Quotation

Product SKU: ASP1000

Reagent Reservoirs

Chemically resistant 15ml reservoirs, 50 per pack

Product SKU: 52001

$22.00

Microwell Plates

Chemically resistant 96-well plates, 10 per pack

Product SKU: 51001

$39.00

mPrep/g™ Pipettor Kit eight-channel

eight-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43200

$686.00

mPrep/g™ Pipettor Kit single-channel

single-channel pipettor, mPrep/g capsule pack, 96 pipette tips

Product SKU: 43100

$351.00