Instruments and Workflows

Cell Culture

cells

Cell culture is one of the major tools used in cellular and molecular biology, providing excellent model systems for studying the effects of drugs, nanoparticles and toxic compounds on cells.

The MWAC Cell culture Facility houses 3 Biosafety cabinets. Two in the general access cell culture room (524D) and one in the secure cell culture room (524A).

For training and access, please contact Dr Joanna Biazik-Richmond.

 

The SEB Cell culture facility is equipped with

2x Class II Biosafety Cabinets 

1x Cytotoxic Hood

Liquid nitrogen storage for cells

Equipment type 

Description

Mammalian cell incubator 

2 x Eppendorf Cell Expert C170i for the purpose of mammalian cell culture (170 L capacity). 

Centrifuge 

1 x Eppendorf 5424 with swinging bucket rotor 

 

1 x Eppendorf 5804 (refrigerated) 

Vortex mixer 

M-SI-100 

Water bath 

NBCT7 (16 L capacity) 

Light microscope 

Leica DM IL (inverted) with ProgRes CFscan 

Biosafety cabinet 

Gelaire Laminar flow cytotoxic drug safety cabinet with Vacusafe system 

 

Nuaire Labgard ES Class II Biosafety Cabinet with Vacusafe system 

Plate reader

CLARIOstar Plus (absorbance, fluorescence, and luminescence)

Biological Sample Preparation

tem

 

Conventional sample preparation for transmission and scanning electron microscopy can be used to study the morphology and ultrastructure of cells to investigate various cellular processes including proliferation, cell death and cell uptake.

For training and access, please contact EMU staff members Dr Joanna Biazik-Richmond, Natasha Kapoor-Kaushik or Greg Harm.

 

The laboratory contains the following biological sample preparation equipment:

1 x Pelco Biowave

2 x Leica Ultramicrotome UC7

1 x Cryo-Ultramicrotome UC6

1 x Reichert Ultramicrotome

Polymerisation Oven

Leica Knife maker

Leica Hot plate

Olympus Microscope with Infinity5 camera

Correlative Light and Electron Microscopy (CLEM)

clem

Correlative Light and Electron Microscopy (CLEM) combines the unique capabilities of light and electron microscopy by studying the exact same sample with both modalities sequentially. The fluorescence signal provides the region of interest and the electron microscopical analysis provides ultrastructural detail and context of the sample. This technique allows us to study rare cellular structures and dynamic cellular processes.

For training and access, please contact BMIF Staff.

 

The light microscopes in the SEB facility include:

1 x Zeiss 800 Confocal with Airyscan (for more details click Zeiss LSM 800)

1 x Label-free Nanolive 3D Cell Explorer

1 x Upright Zeiss confocal LSM 900 with Airyscan 2 equipped with Linkam Cryo-correlative microscopy stage (for more details click Zeiss LSM 900 Cryo)

 

Immunofluorescence and ImmunoEM labelling

cells

For training and access, please contact Dr Joanna Biazik-Richmond or Dr Nicholas Ariotti or Dr Juanfang Ruan.

 

Our facility offers assistance in various distribution analyses by fluorescence and electron microscopy techniques including:

 

-Routine immunofluorescence and CLEM

-Cell transfection and protein expression

-Apex Enzymatic tagging for electron microscopy

-Tokuyasu and and cryosectioning immuno labelling for electron microscopy

-Pre-embedding immuno gold labelling for electron microscopy

 

CryoEM and Cryopreservation

CryoEM and Cryopreservation

Cryofixation is a technique where biological material is rapidly frozen. The speed of the freeze prevents any ice crystal formation and membrane damage from occurring. This form of fixation yields superior preservation thereby achieving a near-to-native state ultrastructure.

For training and access, please contact Dr Nicholas Ariotti or Dr Juanfang Ruan or Dr Joanna Biazik-Richmond.

 

 

The facility is equipped with instruments for cryopreservation including:

-Leica EM GP plunge freezer

-Leica ICE high pressure freezer

-Leica EM AFS2 Automated freeze substitution unit

Volume EM and Electron Tomography

Volume EM and Electron Tomography

Transmission electron tomography and SEM array tomography are two methods of acquiring high resolution volumetric data from cells and organs after resin embedding.

For training and access, please contact Dr Nicholas Ariotti or Natasha Kapoor-Kaushik or Dr Joanna Biazik-Richmond.

 

 

Our facility offers several techniques which are used to generate 3-Dimensional reconstructions of cells and tissue. These advanced techniques can be coupled with correlative fluorescence microscopy, which can be invaluable in understanding how cells and tissue respond to experimental stimuli. Using fluorescent tags allows for the identification and re-registration labelled structures (i.e., nanoparticles) and organelles with both light and electron microscopy.

These techniques include:

-Serial sectioning and scanning electron microscopy (SEM) array tomography.

-Room temperature electron tomography of resin sections

CryoCLEM

CryoCLEM

Cryo-correlative light and electron microscopy is an advanced workflow that allows researchers to use the identifying power of fluorescence microscopy coupled with high-resolution imaging capabilities of the cryo- transmission electron microscope. This methodology facilitates a native and hydrated structural understanding of rare and specific events demarked by laser scanning confocal microscopy with Airyscan detection prior to re-registration of the same region of interest by Cryo-EM or Cryo-Electron Tomography.

For training and access, please contact Dr Nicholas Ariotti.

Plasmid Purification and Transfection

SEB 524 is a PC2 rated and OGTR certified facility (for exempt dealings) which allows the growth of GMOs. For the applications of CLEM and cryo-CLEM specialised plasmids are required for the production of fluorescently tagged markers. These can be transfected into mammalian cell lines and used to tag endocytic pathways or organelles such as, the plasma membrane, the nucleus, the endoplasmic reticulum and the Golgi complex. As part of this workflow we will also need to transform and purify bacterial plasmid vectors.

 

For training and access or for other types of use (e.g microbiology), please contact Natasha Kapoor-Kaushik or Greg Harm.

 

General laboratory equipment list

Includes sample storage in fridge, -30 °C, -80 °C and liquid nitrogen conditions.

 

Equipment type

Description

Bacterial cell incubator

Labec Z-ZWY-103D (capacity)

Centrifuge

Eppendorf 5424R (refrigerated) with swinging bucket rotor

 

Eppendorf 5804

 

Thermofisher Scientific Hergeus Fresco 21

 

Eppendorf minispin

pH meter

Mettler Toledo FiveEasy Plus

Fume hood

2 x Secuflow bench mounted fume hood with services on side walls

Vortex mixer

M-SI-100

Laboratory oven

Labec OEF-65 volume (65 L capacity)

Ultrasonic bath

Unisonics FXP 10DM

Magnetic hot plate

Corning hot plate stirrer PC-420D

 

Labec hotplate stirrer

Rotary suspension mixer

M-TM01600

Electric Bunsen burner

Labec electron Bunsen burner coupled with MC5 controller.

Light microscope

Olympus CKX53 (inverted)

 

Leica M-420 (upright macro)

 

Leica S9E (upright macro)

Spectrophotometer

NanoDrop Lite (for DNA and protein quantification)

Water purification

Milli-Q reference water system (Z00QSV001)

Autoclave service

Location: Science and Engineering Building (E8) level 5 room 515

Contact information: Greg Harm | g.harm@unsw.edu.au | 0437029779 | SEB (E8), level 5, office adjacent to laboratory 524

 

Please read the extended information in the autoclave schedule on the home page.

 

Due to the capacity of the autoclave only one run is required per day (capacity of >25 x 2 L bottles).

 

Weekly schedule:

MONDAY

TUESDAY

WEDNESDAY

THURSDAY

FRIDAY

LIQUIDS CYCLE

(volumes of less than one litre)

LIQUIDS CYCLE (volumes of one to two litres)

+

1 x GOWNS CYCLE (9 am)

DRY CYCLE

LIQUIDS CYCLE (volumes of one to two litres)

Autoclave maintenance and volumes of >2 L (by request)

 

Run times and pick-up times:

Cycle starts at 10:00 am each day. Pick-up will be after 1:15 pm.

 

All that is required is a quick induction and access approval; there is no cost for using the service.