ExploCap™ technology
OZB ExploCap™ RNAs are produced by in vitro transcription and are co-transcriptionnaly capped using trinucleotides ExploCap analog and contain a poly(A) tail at the 3’ end. Sequences have been optimized to yield improved stability and performance.
ExploCap™ is a next-generation co-transcriptional capping reagent designed to generate high-quality capped mRNA with excellent biological properties.
High-quality RNA capping plays a central role in mRNA & saRNA performance, as it directly impacts:
• RNA quality,
• translation efficiency,
• degradation risk by skipping a production step,
• downstream manufacturability.
ExploRNA’s next-generation capping reagent ExploCap™ has been designed to flexibly fit various in vitro transcription (IVT) workflows providing reliable high-quality outcomes. ExploCap™ supports both unmodified and modified RNA, and provides a cost-effective alternative to industry-standard capping reagents.
ExploCap Structure
ExploCap mRNAs resemble fully matured mRNAs with 5’cap structure and 3’ polyA tail, therefore ready to be translated by the ribosome. ExploCap is a proprietary ExploRNA cap with properties similar to Cap 1.
| ARCA | CAP1 | CleanCap® | ExploCap™ | |
|---|---|---|---|---|
| Capping efficiency | 60-80% | 95-99% | 95-99% | 95-99% |
| Transcription Yield | Low | High | High | High |
CleanCap® is a proprietary technology of TriLink Biotechnologies.
ExploCap™ technology is licensed to OZ Biosciences by ExploRNA Therapeutics and is protected under patent WO2021162566.
ExploCap is the intellectual property of ExploRNA Therapeutics.
All RNA products incorporating ExploCap are supplied for Research Use Only (RUO) and are not intended for diagnostic, therapeutic, or clinical use.
Capped mRNAs
ExploCap™ mRNAs are available in unmodified, modified with 5-methoxyuridine (5moU replaces U) or N1-methyl-pseudouridine (N1-mψ) to reduce innate immune responses.
All our messenger RNAs catalog are available capped with ExploCap analog upon request.
ExploCap™ mRNA results
Comparison of transfection efficiency of mRNA capped with Cap1 and ExploCap™ in hek293 cells
Figure 1: HEK293 cultivated in 96-well plates were transfected with 100 ng mRNA encoding for GFP and capped with Cap1 or ExploCap™. 24 H after transfection, GFP expression was visualised under fluorescence microscopie (A) and fluorescence intensity was quantified by flow cytometry (B).
Comparison of transfection efficiency of mRNA capped with Cap1 and ExploCap™ in various cell lines (Vero, C6 and C2C12)
Figure 2: Vero C6 and C2C12 cell lines cultivated in 96-well plates were transfected with 100 ng mRNA encoding for GFP and capped with Cap1 or ExploCap™. 24 H after transfection, GFP expression was visualised under fluorescence microscopie (A) and % of GFP positive cells (B) and fluorescence intensity (C) were quantified by flow cytometry.
Capped saRNAs
ExploCap™ saRNAs are available in unmodified or modified with 5-methyl-Cytidine (m5C) (100% replacement of Cytidine).
All our self-amplifying RNAs catalog are available capped with ExploCap analog upon request.
ExploCap™ saRNA results
Comparison of transfection efficiency of saRNA capped with Cap1 and ExploCap in hek293 cells
Figure 3: HEK293 cultivated in 96-well plates were transfected with 100 ng saRNA encoding for GFP and capped with Cap1 or ExploCap™ using Rmesfect at a 3.5:1 ratio. GFP expression was visualised under fluorescence microscopie at day 1 (D1) and day 6 (D6) after transfection.
Custom mRNA/saRNA Synthesis Service
With over 15 years of nucleic acid delivery expertise, OZ Biosciences is confident in providing you with the best-in-class service for the production of high quality mRNAs & saRNAs.
We are able to synthesis custom messenger RNA (mRNAs) and self-amplifying RNA (saRNAs) at microgram to multigram scales, from a few hundred up to several thousand bases, with a wide variety of modifications including 5’ terminal modifications with cap, internal modification such as 5-methoxyuridine (other modified nucleotides are available) and 3’ modification such as poly-A tail. Fluorescent labeling with Cy5, Cy3 or other options is available.
Production Process
The mRNA production process at OZ Biosciences has been meticulously perfected over the years to achieve an exceptional level of optimization. Using state-of-the-art enzymes, considered to be the most efficient available, the process guarantees production that minimizes dsRNA formation. This attention to detail results in mRNAs of the highest quality, reflecting an ongoing commitment to excellence and innovation in the field.
Quality control
We do know that the reproducibility of scientific experiments is very important to researchers. That is why we developed a quality control system tailored to each product, enabling us to detect defective batches immediately.
Our mRNAs & saRNAs, whether custom or catalog, undergo standard quality control, including measurement of size, integrity and purity by fragment analyzer, agarose gel and Nanodrop (concentration 260/280 and 260/230), a sterility test and an experimental validation (transfection / expression).
However, we have developed additional tests to extend the analysis to include endotoxin detection and double strand RNA (dsRNA) quantification.
| Items | Specification | Standard QC | Superior Grade QC* |
|---|---|---|---|
| Integrity | Agarose gel mobility and fragment analyzer | ✓ | ✓ |
| Concentration | 1mg/ml +/- 5% | ✓ | ✓ |
| A260/280 |
>1.8 for Unmodified mRNAs >1.7 for chemically modified mRNAs |
✓ | ✓ |
| Sterility | Absence of bacterial growth at 37°C | ✓ | ✓ |
| Functionality** | Test for protein expression | ✓ | ✓ |
| Endotoxin | <0.5 EU/mL | ✓ | |
| dsRNA | <0.5% | ✓ |
*Our catalogue mRNAs undergo the standard QC. Superior Grade QC can be performed as an additional prestation. Contact us to get a quote.
** For reporter mRNAs and Spike SARS-Cov2 related mRNAs only for catalog mRNAs. Can be included in superior Grade QC.
Example of Quality Control results :
Agarose gel
Capillary electrophoresis
Endotoxin content
dsRNA content
New enzyme for low dsRNA in our mRNAs!
As part of our continuous quality improvement, we have selected a new generation of RNA polymerase that reduces dsRNA during IVT. We are able to reduce the amount of dsRNA in our catalog and in our customized mRNAs by up to 88%.
Our new quality control limit is 0.05% of dsRNA.
Comparison of dsRNA amount in different mRNA produced by OZ Biosciences before and after the production process improvement.
ExploCap™ GFP mRNA
Unmodified / 5-methoxyuridine / N1-methylpseudouridine
ExploCap™ OVA mRNA
Unmodified / 5-methoxyuridine / N1-methylpseudouridine
ExploCap™ Spike SARS CoV-2...
Unmodified / 5-methoxyuridine / N1-methylpseudouridine
ExploCap™ GFP saRNA
Unmodified / m5C
ExploCap™ F-Luc mRNA
Unmodified / 5-methoxyuridine / N1-methylpseudouridine