WHAT ARE THE ADVANTAGES OF PHOTOPATTERNING WITH PRIMO COMPARE TO OTHER MICROPATTERNING TECHNIQUES?
The PRIMO solution gives researchers complete freedom and full control over their manipulations to optimize the experimental conditions easily. PRIMO allows any shape of protein pattern to be generated rapidly, to precisely align or overlay several proteins (up to 3 without restrictions depending on the proteins used and their interactions) and to control the local density of protein adsorbed to the substrate.
FOR WHICH APPLICATIONS CAN PRIMO BE USED?
- 2D photopatterning on substrate
- Photopatterning on 3D microstructures
- Hydrogel polymerization
HOW LONG DOES IT TAKE TO CREATE A PROTEIN PATTERN WITH THE PRIMO SOLUTION?
The total time to create a pattern (excluding the design step) ranges from 10 to 15 minutes.
The illumination time varies and must be optimized for the patterns projected and the objective used. After each UV illumination and degradation step for the anti-fouling polymer (anti-adhesive), about 5 minutes must be allowed for the proteins to adsorb (time varies with the concentration placed on the substrate).
HOW LONG CAN THE CELL PATTERNS BE STABLE FOR?
The stability period of cell patterns depends on the substrate used and cell type cultured.
Under certain conditions, the cells can adhere to the protein patterns for up to 2 weeks. Our R&D team regularly develops new protocols that are made available to our users.
IS IT POSSIBLE TO PATTERN PROTEINS ON HYDROGELS USING PRIMO?
We recommend using the transfer technique by patterning proteins with PRIMO onto a slide that will be laid on the hydrogel then removed. The UV illumination phase to photopattern proteins onto hydrogels modifies their physical properties and therefore the initially-determined experimental conditions.
CAN THE SLIDE BE COATED IN ADVANCE?
We recommend coating on the same day. However, our R&D team regularly develops new protocols for specific substrates making it possible to keep coated slides for later use.
WHY IS SURFACE TREATMENT IMPORTANT FOR PHOTOPATTERNING?
Photopatterning is a subtractive technology. It first aims to cover a substrate with an anti-fouling polymer that prevents molecules and cells from adhering. The PRIMO system is then used to degrade this anti-adhesive surface locally, under the combined action of UV and the photoactivatable reagent (PLPP).
DOES PRIMO ALLOW EXPERIMENTS TO BE CONDUCTED UNDER STERILE CONDITIONS?
Yes, experiments conducted with PRIMO can be carried out under sterile conditions, even if the PRIMO device is not set up in a room assigned to this purpose. In such cases, we recommend performing all rinsing and incubation steps in a biosafety cabinet (BSC), then putting the sample in a closed culture plate that can then be placed on the motorized stage of the PRIMO photo-patterning device.
CAN PRIMO BE USED TO PATTERN PROTEINS ON 3D STRUCTURES?
PRIMO can be used to pattern all surfaces, whether flat, curved or microstructured. For example, it is possible to pattern proteins on microwell walls or PDMS micropillars. In-house data available on request.
DOES THE LIGHT-ACTIVATED PLPP REAGENT ALWAYS HAVE TO BE USED WITH PRIMO?
The PLPP accelerates the degradation reaction of the anti-fouling polymer during photopatterning (a few seconds instead of a few hours). However, microfabrication manipulations with SU8 resins or NOA do not require the use of PLPP, since these products are photosensitive in the PRIMO wavelength (365 nm for the PRIMO 2 system, previously 375 nm).
WHAT ANTI-FOULING SURFACE TREATMENT CAN BE CARRIED OUT ON THE SUBSTRATES?
We recommend using PLL-PEG. We regularly develop new protocols to enhance the stability of surface coatings.
WHAT IS THE DEPTH OF FIELD FOR THE DEVICE?
By using a 20x objective, the depth of field for the laser passing through a transparent substrate is 200 µm to 300 µm.
WHAT IS THE RESOLUTION OF THE DEVICE?
The resolution is 1.2 µm over the entire field of view of approximately 500x300 µm with a 20x objective.
WHAT IS THE MAXIMUM SIZE THAT CAN BE PATTERNED?
PRIMO allows all desired images to be patterned, without size limitation.
WHICH PROTEINS CAN BE PATTERNED WITH THE PRIMO DEVICE?
Our team and users employ more than ten different proteins, including Fibrinogen-488, Fibrinogen‐647, Fibronectin, GFP, Neutravidin‐488, Neutravidin‐647, PLL‐PEG‐Biotin, Protein A‐647 and Streptavidin, as well as primary and secondary antibodies.
WHAT SUBSTRATES CAN BE USED FOR PROTEIN PHOTO-PATTERNING WITH PRIMO?
PRIMO can be used to pattern all standard cell culture substrates: PDMS, glass, plastic, NOA,
SU8. New substrates are regularly tested and validated by our in-house R&D team. In-house
data available on request.
WHAT TYPES OF PHOTORESIST CAN BE USED FOR MICROFABRICATION WITH PRIMO?
All photoresist with a sensitivity between 365 and 375 nm can be used with PRIMO. Our R&D team mainly works with SU8 photoresists.