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mPrep™ ASP-1000 and ASP-2000 Specimen Processing Speeds Compared to Other Methods

This page provides examples that demonstrate how mPrep ASP processors accelerate preparation speed and reduce hands-on effort in comparison to other methods.  Of course, tissue preparation speeds for TEM and volume EM (vEM) vary due to the specimen type, staining, how the specimens were prepared before processing at the electron microscope lab, and due to different routines practiced in different labs.

TEM tissue preparation using ASP-1000 and ASP-2000 processors.

Specimens

Protocol Summary

Process Time

Source/Citation

Kidney (Rat). Eight 8 2mm cube specimens

ASP: Glutaraldehyde-Paraformaldehyde (GAPFA) rinse-out, OsO4, Ethanols, Acetone, 100% epoxy infiltration before curing

46 minutes

Stewart et al. (2017) Fast, Walk-away, Automated Processing of Mammalian Tissue for LM and TEM. Microsc. Microanal. PDP52.

Kidney, Human, 18G clinical biopsies. 1-16 patient specimens consisting of up to eight 2mm long tissue pieces: Total up to 128 biopsy segments.

ASP: GAPFA rinse-out, OsO4, Ethanols, Acetone, 100% epoxy infiltration, transfer to flat mold for curing.

3 hours 20 minutes

Flint N & Goodman SL (2023) An Efficient Clinical Pathology TEM Workflow from Specimen Acquisition Through Automated Preparation and Electron Microscopy. National Society for Histotechnology 2023, Baltimore, W67.

Planarian worm, Schmidtea mediterranea

ASP: GAPFA rinse- out, OsO4, Ethanols, Acetone, 100% epoxy infiltration.

5 hours

McClain M (2016) Bringing Speed and Automation Together with the ASP-1000. Microsc Microanal. PDP49.

Liver (rat), Extraocular muscle sarcomeres (mouse), Nerves – neuronal somas from dorsal root ganglion (mouse). Eight  1-2mm cubes

ASP: GAPFA rinse out, no OsO4, ethanols, through 100% LR White acrylic infiltration

45 minutes

Stewart NR et al. (2017) Preparing LR White Embedded Tissue with mPrep/s Specimen Capsules. Microsc. Microanal. PDP56.

Kidney, Liver, Heart, Skeletal muscle (Rat). Eight 1-2mm cube specimens

ASP: GAPFA rinse out, OsO4, Ethanols, Acetone, 100% epoxy infiltration

2 hours 12 minutes

Stewart NR et al. (2017) Fast, Walk-away, Automated Processing of Mammalian Tissue for LM and TEM. Microsc. Microanal. PDP52.

Skeletal muscle (rat)

ASP: GAPFA rinse out, OsO4, Ethanols, Acetone, 100% epoxy infiltration

2 hours 12 minutes

Yeast, agar enrobed pellet

ASP: GAPFA rinse-out, OsO4, Ethanols, Acetone, 100% epoxy infiltration.

1 hour 30 minutes

Tetri et al.  (2018). Sex-Specific Skeletal Muscle Fatigability and Decreased Mitochondrial Oxidative Capacity in Adult Rats Exposed to Postnatal Hyperoxia.  Front. Physiol. doi: 10.3389/fphys.2018.00326.

Kidney (rat). Eight 1.02mm diameter needle biopsies

Agar enrobe GAPFA fixed biopsy, then ASP: GAPA rinse out, OsO4, Uranyl acetate (UA), Ethanols, Acetone, 100% epoxy infiltration.

2 hours

Strader, T., & August, B. (2019). Automated Preparation of Core Needle Biopsy Specimens for TEM Imaging. Microsc Microanal. 25(S2), 1184-1185.

TEM tissue preparation using manual, microwave oven and carousel-type tissue processors.

Specimens

Protocol Summary

Process Time

Source/Citation

Most biological tissues and specimens

Manual: GAPFA or glutaraldehyde, aldehyde rinse-out, OsO4, graded ethanols and/or acetones, resin infiltration.

Manual: 18 hours to 1 day 12 hours before resin curing

Electron Microscopy, 2nd Edition, JJ Bozzola and LD Russell, Jones and Bartlett, Boston, 1999.

Most biological tissues and specimens

Manual: GAPFA or glutaraldehyde, aldehyde rinse-out, OsO4, graded ethanols and/or acetones, resin infiltration.

Manual: 12 hours to 22 hours before resin curing

Methods in Molecular Biology, vol. 369 Electron Microscopy: Methods and Protocols, Second Edition Edited by: J. Kuo © Humana Press Inc., Totowa, NJ.

Placenta (goat), Liver (mouse), kidney (mouse), esophagus (deer)

Manual: GAPFA, OsO4, Acetones, resin infiltration, then 24 hr resin cure.

Microwave: GAPFA, OsO4, Acetones, resin infiltration, then 24 hr resin cure.

Manual: 1 day 11 hours before resin curing.

Microwave: 1 hour 3 minutes before resin curing.

Giberson RT et al (1997) Four-hour processing of clinical/diagnostic specimens for electron microscopy using microwave technique. J Vet Diagn Invest 9:61-67.

Kidney, cilia, thyroid

Microwave: GAPFA, GAPFA rinse-out, OsO4, ethanol and/or Acetones, epoxy resin infiltration

Microwave: 2 hours including epoxy resin curing.

Giberson RT et al (2003) Microwave and Digital Imaging Technology Reduce Turnaround Times for Diagnostic Electron Microscopy. Ultrastructural Pathology, 27:187–196.

Cell suspensions, Liver (human and mouse), C Elegans

Microwave: Aldehyde fix rinse-out, OsO4, ethanols, acetone, epoxy infiltration

Carousel tissue processor: Aldehyde fix rinse-out, OsO4, ethanols, acetone, epoxy infiltration

Microwave: 3 hours 20 minutes to 4 hours 25 minutes, including 1 hour 45 minutes epoxy polymerization

Carousel tissue processor: 3 days 2 hours 25 minutes, including 2 days epoxy polymerization

Schroeder JA et al. (2006) Microwave-assisted tissue processing for same-day EM-diagnosis of potential bioterrorism and clinical samples. Micron 37:577–590.

Retina (mouse)

Manual: GAPFA, OsO4, Uranyl acetate (UA) Acetones, epoxy resin infiltration

Microwave: GAPFA, OsO4, Uranyl acetate (UA) Acetones, epoxy resin infiltration

Manual: 2 days 23 hours before epoxy resin curing.

Microwave: 2 hours before epoxy resin curing.

Wendt KD et al (2004) Comparison of conventional and microwave-assisted processing of mouse retinas for transmission electron microscopy. J.  Microscopy 214(1):80 – 88

vEM specimen preparation using ASP-1000 and ASP-2000 processors and manual preparation

Specimens

Protocol Summary

Process Time

Source/Citation

Brain (rat)

ASP: GAPFA rinse- out, tannic acid (TA), Os-KFeCN, Thicarbohydrazide (TCH), OsO4, UA, Pb, ethanols, acetone, 100% epoxy- resin infiltration

Manual: GAPFA rinse- out, tannic acid (TA), Os-KFeCN, Thicarbohydrazide (TCH), OsO4, UA, Pb, ethanols, acetone, 100% epoxy- resin infiltration

ASP: 8 hours ASP processing, then overnight epoxy resin cure

Manual: 4 days and 2 overnights, including epoxy cure

Benson EK et al. (2020) Serial Block-Face SEM of Brain Tissue Using Rapid Automated Preparation. Microsc. Microanal. 26 (Suppl 2): 1372.

Brain, Liver (rat)

ASP: GAPFA rinse- out, tannic acid (TA), Os-KFeCN, Thicarbohydrazide (TCH), OsO4, UA, Pb, ethanols, acetone, 100% epoxy- resin infiltration

Manual: GAPFA rinse- out, tannic acid (TA), Os-KFeCN, Thicarbohydrazide (TCH), OsO4, UA, Pb, ethanols, acetone, 100% epoxy- resin infiltration

ASP: 8 hours ASP processing, then overnight epoxy resin cure. 1 hour hands-on effort

Manual: 4 days, then 2 days epoxy cure.  2 days effort

Goodman SL et al. (2019) Rapid Automated Preparation for Serial Block Face Scanning Electron Microscopy. Microsc. Microanal. 25(Suppl 2):1182.

Flatworm (Schmidtea mediterranea)

ASP: Aldehyde rinse-out, Os-KFeCN, TCH, OsO4, UA, Pb, ethanols, acetone, 100% epoxy infiltration

19 hours ASP processing, then 2 days epoxy resin cure

McClain M et al. (2020) Automated Heavy Metal Tissue Staining for Serial Block Face Imaging with the ASP-1000. Micro Microanal PDP52.

Heart (rat)

ASP: GAPFA rinse-out, Os-KFeCN, TCH, OsO4, UA, Pb, ethanols, acetone, 100% epoxy infiltration

7 hours 30 minutes ASP processing, then overnight epoxy resin cure

Goodman SL et al. (2019) Effects of Postnatal Hyperoxia Exposure on Cardiac Muscle Examined with 3D Serial Block Face SEM using Rapid Automated Sample Preparation . Micro Microanal. 25:PDP46.

Brain (marmoset) and Breast cancer tumors (mouse)

ASP: GAPFA rinse-out, Os-KFeCN, TCH, OsO4, UA, Pb, ethanols, acetone, 100% epoxy infiltration

Manual: GAPFA rinse-out, Os-KFeCN, TCH, OsO4, UA, Pb, ethanols, acetone, 100% epoxy infiltration

ASP: 7 hours.  2.5 hours active technician time.

Manual: 2.5 days.  5.5 hours active technician time.

Stempinski, E. et al. (2023). Chapter 1 – Automatic Large Volume Sample Preparation for vEM. Methods in Cell Biology. 177, 1-32.

Planarian – Schmidtea mediterranea

ASP: GAPFA rinse-out, reduced OsO4, TCH, OsO4, UA, Pb, acetone, 100% epoxy infiltration

Manual: GAPFA rinse-out, reduced OsO4, TCH, OsO4, UA, Pb, acetone, 100% epoxy infiltration

Process times not provided for ASP preparation.  Publication states:

“We developed this protocol to run both manually on the bench and with an automated sample processing robot (ASP1000, Microscopy Innovations). The sample processing robot has the advantage of being almost completely hands-off once set up, as well as far faster, given that the robot has continuous rapid agitation and runs after working hours.”

McClain ML and Nowotarski SH (2023). Chapter 9 – Serial block-face scanning electron microscopy of Schmidtea mediterranea. Methods in Cell Biology. 177, 1-32.

Breast cancer tumors (mouse)

ASP: GAPFA rinse-out, Os-KFeCN, TCH, OsO4, UA, Pb, ethanols, acetone, 100% epoxy infiltration

Manual: GAPFA rinse-out, Os-KFeCN, TCH, OsO4, UA, Pb, ethanols, acetone, 100% epoxy infiltration

ASP: 5 hours.  2.5 hours active technician time.

Manual: 20 hours.  5.5 hours active technician time.

Stempinski, E. et al. (2022). Automatic Sample Processing for vEM in a Mouse Model of Breast Cancer. Microsc Microanal. 28(S1), 1370.

Brain

Manual: GAPFA rinse-out, Os-KFeCN, TCH, OsO4, UA, Pb aspartate, ethanols, acetone, 100% epoxy infiltration

3-4 days processing, and 2-3 days resin cure

Hua, Y., Laserstein, P. & Helmstaedter, M. (2015) Large-volume en-bloc staining for electron microscopy-based connectomics. Nat Commun. 6:7923.

Mammalian tissues and others

Manual: GAFPA rinse-out, Os-KFeCN, TCH, OsO4, UA, Pb aspartate, ethanols, acetone, 100% epoxy infiltration

2-4 days processing, and 2 days resin cure

Deerinck TJ et al. (undated) Preparation of Biological Tissues for Serial Blockface Scanning EM (SBEM).