TECHNOLOGY EXPLAINER
mRNA Medicines and Lipid Nanoparticles
— the nucleic acid is the star, but most of the mass is lipid and sugar
An mRNA medicine is often described as "a drug that delivers genetic instructions", yet weigh what is actually in the vial and the mRNA is only a small fraction. A single dose contains about 25 times as much lipid as mRNA, and about 1,000 times as much sugar. In practice the technology is a combination of nucleic acid chemistry, lipid formulation design and a mixing process.
- What an mRNA medicine is (the short version)
- The structure of mRNA — five parts
- Why modified nucleosides are needed
- How it is made — from template DNA to IVT
- A materials engineer's view (1): removing impurities pays off by orders of magnitude
- Lipid nanoparticles (LNPs) — what the four lipids do
- How LNPs are made — the rapid-mixing step
- Our calculation: how many milligrams of what go into one dose
- A materials engineer's view (2): an LNP is a material defined by its formulation and mixing conditions
- How quality is measured — what the specifications tell you
- The cold chain — why storage temperatures differ between products
- Glossary / References / Claim-to-source audit
Sourced = stated in published material (link given)
Our calculation = a figure this article derived, with the assumptions spelled out
Not yet confirmed = a plan or a target with no confirmed track record
Structural summaries and readings from a materials or process-design standpoint are marked separately as Commentary.
This article explains materials and manufacturing technology. It does not assess the efficacy or safety of any treatment or vaccine, and it is not medical advice. Approval status and product composition are quoted only where they appear in material published by the regulators (the FDA and the EMA).
1. What an mRNA medicine is (the short version)
Inside a cell, the information in DNA is copied into mRNA, and ribosomes use that copy to build proteins. The idea behind an mRNA medicine is to deliver only that working copy of the blueprint from outside.
- What is delivered: Pardi and colleagues describe mRNA as the intermediate step between the translation of protein-encoding DNA and the production of proteins by ribosomes in the cytoplasmSourced. Two kinds of RNA are being studied as vaccines: non-replicating mRNA and virally derived self-amplifying RNASourced
- What is said in its favour: the same review states that because mRNA is a non-infectious, non-integrating platform, there is no potential risk of infection or insertional mutagenesisSourced
- What makes it hard: left as it is, mRNA is broken down, and the immune system detects it as foreign. Modified nucleosides and lipid nanoparticles are what solved those two problemsSourced
Think of an mRNA medicine only as "a nucleic acid drug" and you will misread what manufacturing it involves. Working from the composition in the FDA prescribing information, one dose (the 30 µg presentation) contains about 0.76 mg of lipid in total — about 25 times the mass of the mRNAOur calculation. Physically, the product is a lipid dispersion with mRNA inside it, and its performance depends heavily on the lipid ratios and the mixing conditions.
2. The structure of mRNA — five parts
Hou and colleagues set out the structural elements of mRNA as followsSourced.
mRNA typically contains five structural elements: a 5′ cap, a 3′ poly(A) tail, the protein-coding sequence, and the 5′ and 3′ untranslated regions (UTRs).Sourced
The paper describes these elements as critical to the initiation, translation, termination, post-transcriptional modification and degradation of the mRNA molecule, and notes that each can be engineered to improve stability and translation efficiencySourced.
3. Why modified nucleosides are needed
The 2023 Nobel Prize in Physiology or Medicine went to the research that answered this question.
The Nobel Assembly at Karolinska Institutet announced that it had decided to award the 2023 Nobel Prize in Physiology or Medicine jointly to Katalin Karikó and Drew Weissman for their discoveries concerning nucleoside base modifications that enabled the development of effective mRNA vaccines against COVID-19.Sourced
The announcement explains how they got there. The two noticed that dendritic cells recognise in vitro transcribed mRNA as a foreign substance, which leads to their activation and the release of inflammatory signalling molecules, and focused on the fact that the mRNA of mammalian cells carries base modifications, whereas in vitro transcribed RNA does notSourced.
The result was clear-cut: when base modifications were included, the inflammatory response was almost abolished. The work was published in 2005Sourced.
The abstract of the original 2005 paper (Karikó et al., Immunity) says the followingSourced.
RNA signals through human TLR3, TLR7 and TLR8, but incorporating the modified nucleosides m5C, m6A, m5U, s2U or pseudouridine abolishes that activity.Sourced
Its conclusion was that nucleoside modification suppresses the ability of RNA to activate dendritic cellsSourced.
Later, N1-methylpseudouridine (m1Ψ) was reported to do even better. In the abstract of their 2015 paper, Andries and colleagues report that mRNA carrying the m1Ψ modification, alone or in combination with m5C, outperformed the then-standard pseudouridine (Ψ) and/or m5C/Ψ-modified mRNA platforms, with reporter gene expression up to about 44 times higher on transfection into cell lines (double-modified versus double-modified) and about 13 times higher in mice (single-modified versus single-modified)Sourced.
The EMA's public assessment report states, for the active substance of Comirnaty, that the RNA contains no uridine at all; modified N1-methylpseudouridine is used in its place during RNA synthesisSourced.
The FDA prescribing information likewise names the active ingredient as nucleoside-modified messenger RNA (modRNA)Sourced.
4. How it is made — from template DNA to IVT
Making an mRNA medicine involves no cell culture at all. That is the decisive difference from viral vectors and antibody drugs.
Preparing a linearised DNA template is the first step in GMP manufacture of mRNA. From that template, mRNA is transcribed in vitro in the presence of an RNA polymerase and ribonucleoside triphosphates. The remaining DNA template is removed by DNase digestion, and the mRNA is capped by chemical or enzymatic methods. Finally, the mRNA is purified by microbead-based precipitation or by chromatography to remove enzymes, free nucleotides, truncated nucleic acid fragments and double-stranded RNA. The purified mRNA can be dissolved in a storage buffer, sterile-filtered and frozen for long-term storage.Sourced
The EMA public assessment report describes the same skeleton for the active substance of Comirnaty: the manufacturing process for the BNT162b2 active substance consists of five main steps. The RNA is synthesised from linear DNA by an in vitro transcription (IVT) step, which is followed by several purification and filtration steps. Finally, the RNA undergoes a final filtration before being dispensed and stored frozenSourced.
On where the template comes from, the report says: the BNT162b2 active substance is made by in vitro transcription from a linear DNA template, which is produced via plasmid DNA obtained from transformed E. coli cells. The linear DNA template is not part of the final product, but because it defines the sequence of the mRNA product it is fundamentally important to ensuring proper control of the active substanceSourced.
On the enzymes used for in vitro transcription, Pardi and colleagues state that IVT mRNA is produced from a linear DNA template using T7, T3 or SP6 phage RNA polymeraseSourced. The 5′ cap can be added during or after the transcription reaction, either by the vaccinia virus capping enzyme or by incorporating synthetic cap analogues or anti-reverse cap analogues, and the poly(A) tail is added either directly from the encoding DNA template or by using poly(A) polymeraseSourced.
5. A materials engineer's view (1): removing impurities pays off by orders of magnitude
IVT is an enzymatic reaction, but it does not produce only the target molecule. Pardi and colleagues put it this way:
enzymatically synthesised mRNA preparations contain double-stranded RNA (dsRNA) contaminants as aberrant products of the IVT reactionSourced.
What makes this awkward is that dsRNA is not "just an impurity": it is the signal that switches on the cell's defence machinery. The same paper explains that recognition of dsRNA leads to strong type I interferon production, raises the expression and activation of PKR and OAS, and ends in inhibition of translation and degradation of cellular mRNA and ribosomal RNASourced. The impurity stops the product itself from working.
That is why removing it has such an extreme effect. The paper states that purification by FPLC has been shown to increase protein production from IVT mRNA in primary human dendritic cells by up to 1,000-foldSourced.
Remove a few per cent of impurity and performance rises a thousandfold — in the materials world, the only things with that kind of sensitivity are metallic impurities in semiconductor materials and catalyst poisons (our commentary). In the sense that the purity specification is not a nice-to-have but a condition for the product to work at all, the design mindset required is closer to that for high-purity materials than for ordinary functional materials.
Indeed, the EMA public assessment report lists among the release tests for the active substance RNA integrity (capillary gel electrophoresis), 5′ cap (RP-HPLC), poly(A) tail (ddPCR), residual DNA template (qPCR) and dsRNA (immunoblot)Sourced. That dsRNA has a specification item of its own shows how much weight this impurity carries (our commentary).
6. Lipid nanoparticles (LNPs) — what the four lipids do
Purified mRNA does not work in the body on its own: it is broken down, and it cannot get inside cells. So it is packaged in lipid nanoparticles (LNPs). Hou and colleagues explain that LNP–mRNA formulations show a stable nanostructure, with the mRNA molecules encapsulated in the inner core through electrostatic interactions with the lipids, and that this structure protects the mRNA from degradation by nucleases and improves the stability of the particles in physiological fluidsSourced.
An LNP is a blend of four kinds of lipid, each with a different job.
| Component | Role (as described in the paper) | Comirnaty | Spikevax |
|---|---|---|---|
| Ionisable lipid | Protonated and positively charged at low pH but neutral at physiological pH. Being neutral reduces interactions with the anionic membranes of blood cells and improves biocompatibility. Protonated at the low pH inside endosomes, it can promote membrane destabilisation and endosomal escape | ALC-0315 | SM-102 |
| Helper lipid (structural lipid) | DSPC is a phosphatidylcholine with saturated acyl chains, a melting point of about 54 °C and a cylindrical molecular shape, so it forms a lamellar phase and stabilises the particle structure | DSPC | DSPC |
| Cholesterol | Can improve particle stability by modulating membrane integrity and rigidity | Cholesterol | Cholesterol |
| PEG lipid | The amount changes particle size and zeta potential. It limits particle aggregation and contributes to stability, and certain modifications reduce clearance by the kidneys and the mononuclear phagocyte system, extending circulation time | ALC-0159 | PEG2000-DMG |
All role descriptions are Sourced (Hou et al. 2021 [Reference 4]). Component names for the products follow the FDA prescribing information [References 5 and 7] and the EMA public assessment report [Reference 6]. The EMA classes ALC-0315 and ALC-0159 as novel excipients and notes that DSPC and cholesterol are already used in several approved productsSourced.
7. How LNPs are made — the rapid-mixing step
The way LNPs are made has changed a great deal over the past twenty years or so.
Historically, formulations of lipid nanoparticles with nucleic acids were produced by methods such as thin-film hydration and reverse-phase evaporation, and the particle size was then made more uniform by extrusion.Sourced
Lipid nanoparticle–mRNA formulations are now generally produced by rapid mixing, in which an ethanol phase (the lipid components) and an aqueous phase (the mRNA molecules) are mixed under specific conditions, namely pH and flow rate. This technique allows lipid nanoparticle–mRNA formulations with high encapsulation efficiency and a uniform size distribution to be produced reproducibly and at scale.Sourced
The leading example is microfluidic mixing. In the abstract of their 2015 paper, Leung and colleagues note that earlier work had shown that lipid nanoparticles (LNPs) composed of an ionisable cationic lipid, a PEG lipid, distearoylphosphatidylcholine (DSPC), cholesterol and siRNA can be made efficiently using microfluidic mixing, and report that the same method extends to encapsulating polyanions larger than siRNA, namely mRNA (1.7 kb) and plasmid DNA (6 kb)Sourced.
The paper also finds that as the proportion of bilayer-forming DSPC is raised, LNP–siRNA systems take on a more bilayer-like structure, and states that the core of LNP–siRNA systems can adopt a continuum of nanostructures depending on the proportions of the constituent lipids and their structural preferencesSourced.
8. Our calculation: how many milligrams of what go into one dose
Composition of Comirnaty (2026-2027 Formula, 0.3 mL, 30 µg presentation)Sourced
- Nucleoside-modified mRNA: 30 µg (= 0.030 mg)
- ALC-0315 (ionisable lipid): 0.43 mg
- ALC-0159 (PEG lipid): 0.05 mg
- DSPC: 0.09 mg / cholesterol: 0.19 mg
- Tromethamine: 0.06 mg / tromethamine hydrochloride: 0.4 mg / sucrose: 31 mg
Composition of Spikevax (2026-2027 Formula, 0.5 mL, 50 µg presentation)Sourced
- Nucleoside-modified mRNA: 50 µg (= 0.050 mg)
- Total lipids: 1.01 mg (SM-102, PEG2000-DMG, cholesterol, DSPC)
- Tromethamine 0.25 mg / tromethamine hydrochloride 1.2 mg / acetic acid 0.021 mg / sodium acetate trihydrate 0.10 mg / sucrose: 43.5 mg
Converting the figures aboveOur calculation:
- Total lipid in Comirnaty: 0.43 + 0.05 + 0.09 + 0.19 = 0.76 mg
- Lipid to mRNA by mass: 0.76 ÷ 0.030 = about 25 times
- Lipid composition by mass: ALC-0315 about 57% / cholesterol 25% / DSPC about 12% / ALC-0159 about 7%
- Sucrose to mRNA by mass: 31 ÷ 0.030 = about 1,000 times
- Lipid to mRNA in Spikevax: 1.01 ÷ 0.050 = about 20 times; sucrose to lipid is 43.5 ÷ 1.01 = about 43 times
- The ratio holds across presentations: in the 10 µg Comirnaty presentation (for ages 5 to 11) total lipid is 0.14 + 0.02 + 0.03 + 0.06 = 0.25 mg, and 0.25 ÷ 0.010 = about 25 times — the same ratio as the 30 µg presentation
Assumptions and limits: the lipid total is simply the sum of the individual values stated on the label; it does not show the input ratios (molar ratios) used in manufacture. The Spikevax label gives only total lipids, so no per-lipid ratios are calculated for it. These figures are also meant only to look at the formulation from a materials standpoint; they say nothing about efficacy or safety.
9. A materials engineer's view (2): an LNP is a material defined by its formulation and mixing conditions
What the calculation in Section 8 shows is that an LNP is not merely "a container for the active ingredient"; it is a formulated material in its own right. Ionisable lipid at 57% of the mass, cholesterol at 25%, DSPC at 12% and PEG lipid at 7%Our calculation — these proportions can be read exactly the way you would read the formulation sheet of any functional material.
- Main ingredient: the ionisable lipid. Its pH responsiveness — positively charged at low pH, neutral at physiological pH — is the core of the designSourced
- Framework: DSPC. With a melting point of about 54 °C and a cylindrical shape, it forms a lamellar phaseSourced. Another phospholipid, DOPE, has a melting point of about 30 °C and a conical shape and readily adopts an inverted hexagonal phase, so its character is quite differentSourced
- Stiffness control: cholesterol, which modulates membrane integrity and rigiditySourced
- Surface modification: the PEG lipid. The amount changes particle size and zeta potential and limits aggregationSourced
Note how molecular shape and phase behaviour directly set the structure of the formulation. Cylindrical means lamellar, conical means inverted hexagonal — this is exactly the critical packing parameter familiar from surfactant science (our commentary). It is the same point Leung and colleagues make when they state that the core of LNP–siRNA systems can adopt a continuum of nanostructures depending on the proportions of the constituent lipids and their structural preferencesSourced.
And the formulation alone does not settle it. Hou and colleagues explicitly name specific conditions, namely pH and flow rate, for rapid mixingSourced. Same formulation, different mixing, different particle — a feeling anyone who has worked with emulsification or nanoparticle dispersions will recognise (our commentary).
The EMA public assessment report contains a passage worth reading from a materials-sourcing standpoint: the functional lipid excipients ALC-0315 and ALC-0159 are classed as novel excipients, while the structural lipids DSPC and cholesterol are already used in several approved productsSourced.
For DSPC, the report goes on to explain why it is not regarded as a novel excipient: DSPC is used as part of the LNP in Onpattro, a product approved in the EU, and Onpattro is given intravenously at doses far higher than the intramuscular dose of this productSourced.
A material with a track record and one without require different amounts of documentation at review. In other words, material selection is constrained not only by performance but by whether a precedent exists (our commentary). Bringing in a new material is itself booked as a development cost, and for anyone supplying materials to a regulated industry that is information as important as the performance specification.
On Onpattro, Hou and colleagues state that the ionisable lipid DLin-MC3-DMA (MC3) is a key delivery component of Onpattro, the first siRNA drug approved by the FDASourced. A lineage running from MC3 to SM-102 and ALC-0315 can therefore be traced in published material. Hou and colleagues describe a design trend of introducing ester bonds to make the lipids more biodegradable, and summarise that SM-102 and ALC-0315 show better in vivo delivery efficiency and pharmacokinetics than MC3Sourced.
10. How quality is measured — what the specifications tell you
The EMA public assessment report lists the specification items for release and stability testing of the finished product. What is measured tells you what is fragile (the grouping is this article's own).
| What is measured | Method (as stated in the EPAR) | What it checks (our reading) |
|---|---|---|
| LNP particle size | Dynamic light scattering (DLS) | Whether the particles have formed at the intended size |
| LNP polydispersity | Dynamic light scattering (DLS) | The spread of particle sizes, which reflects directly how reproducible the mixing conditions are |
| RNA encapsulation | Fluorescence assay | Whether the mRNA is inside the particles rather than leaking out |
| RNA content | Fluorescence assay | The amount of active ingredient |
| Content of each lipid | HPLC-CAD | The amount of each of the four components, i.e. whether the formulation ratio has been held |
| Lipid identity | HPLC-CAD | Whether the lipids present are the right ones |
| RNA integrity | Capillary gel electrophoresis | Whether the mRNA has been broken |
| Potency / in vitro expression | Cell-based flow cytometry | Whether protein is actually made |
| Appearance, pH, osmolality, subvisible particles, endotoxin, sterility and others | European Pharmacopoeia methods and others | Basic requirements for an injectable |
All Sourced (EMA public assessment report [Reference 6]). The active-substance specification includes RNA integrity, 5′ cap, poly(A) tail, residual DNA template and dsRNASourced.
Particle size, polydispersity, encapsulation and the content of each lipid — the fact that these four appear in the release specification means they can shift with process conditions (our commentary). The EMA report itself records that concerns were raised during the procedure about the acceptance criteria for LNP size, polydispersity, RNA encapsulation, in vitro expression and RNA integrity, and that the criteria were tightenedSourced. For a new material system, the specifications themselves settle while still moving — and here that process is preserved in the public record.
11. The cold chain — why storage temperatures differ between products
It is widely known that mRNA–LNP products need cold transport, but the conditions differ from product to product, and they have changed over time.
| Product | Frozen storage | Refrigerated handling |
|---|---|---|
| Comirnaty (single-dose vials) | In an ultra-low-temperature freezer at −90 °C to −60 °C. Must not be stored at −25 °C to −15 °C | If moved to 2 to 8 °C immediately on receipt, can be kept for up to 10 weeks. Must not be refrozen after thawing. Total time at 8 to 25 °C must not exceed 12 hours |
| Comirnaty (prefilled syringes) | Do not freeze | Store at 2 to 8 °C. Total time at 8 to 25 °C no more than 12 hours |
| Spikevax | Stored frozen at −50 °C to −15 °C | After thawing, at 2 to 8 °C for up to 60 days (or until the printed expiry date, whichever comes first) |
All Sourced (FDA prescribing information [References 5 and 7]). On transport of Comirnaty, the EMA public assessment report states that the transport temperature range of −90 to −60 °C is based on the available stability data and that one thaw and refreeze cycle is permitted during transportSourced.
As background to these differences, Hou and colleagues point out that the storage form (aqueous, frozen or lyophilised) and the type of cryoprotectant (sucrose, trehalose or mannitol) affect long-term stability, and state that the approved COVID-19 mRNA vaccines are all stored frozen in the presence of sucroseSourced. They also cite a report that when 5% (w/v) sucrose or trehalose was added and the formulation stored in liquid nitrogen, mRNA delivery efficiency in vivo was maintained for at least three monthsSourced.
The 31 mg of sucrose (about 1,000 times the mRNA)Our calculation that came out of the calculation in Section 8 takes on a different meaning in this context. The cryoprotectant is the heaviest component in the formulation, and it decides whether the product can survive the process of freezing and thawing (our commentary).
Hou and colleagues state that cold-chain transport can preserve vaccine activity, but developing lipid nanoparticle–mRNA formulations that do not need cold or frozen storage would not only lower manufacturing and transport costs but also speed up wider accessSourced. How far this problem had been solved as of September 2026 is not stated here, because no primary source confirming it could be found within the scope of this article.
Approved products (as confirmed in material published by the regulator)
| Product | What the regulator states | First approval |
|---|---|---|
| COMIRNATY | FDA: STN 125742, proper name "COVID-19 Vaccine, mRNA", manufacturer BioNTech Manufacturing GmbH. A sterile suspension for intramuscular injection whose active ingredient is nucleoside-modified messenger RNA (modRNA) | 23 August 2021 (earliest approval letter) |
| SPIKEVAX | FDA: STN 125752, proper name "COVID-19 Vaccine, mRNA", manufacturer Moderna Tx, Inc. A sterile suspension for intramuscular injection whose active ingredient is nucleoside-modified messenger RNA (mRNA) | 31 January 2022 (earliest approval letter) |
All Sourced (FDA product pages [References 10 and 11] and prescribing information [References 5 and 7]). This article does not cover the indications or clinical results of these products. Note that the Nobel announcement states that protective efficacy of around 95% was reported and that both vaccines were approved as early as December 2020Sourced (this is the announcement's statement, not this article's assessment).
- mRNA is made of five parts: the 5′ cap, 5′ UTR, coding region, 3′ UTR and poly(A) tailSourced
- Replacing uridine with N1-methylpseudouridine was the key to practical use. It was the subject of the 2023 Nobel Prize in Physiology or MedicineSourced
- Manufacture needs no cell culture. It is a chain of chemical and enzymatic steps: template DNA, IVT, DNase digestion, capping, purificationSourced
- Removing the impurity (dsRNA) pays off by orders of magnitude. FPLC purification has been reported to raise protein production by up to 1,000-foldSourced
- An LNP is a four-lipid formulated material. By mass: ionisable lipid about 57%, cholesterol 25%, DSPC about 12%, PEG lipid about 7%Our calculation
- Most of the mass of the product is not mRNA. There is about 25 times as much lipid and about 1,000 times as much sucroseOur calculation
12. Glossary
- mRNA
- Messenger RNA. A single-stranded nucleic acid that carries the blueprint for a protein.
- IVT (in vitro transcription)
- A reaction that synthesises RNA from a DNA template in a test tube, without using cells.
- Template DNA
- The linear DNA that IVT copies from, made from plasmid DNA grown in E. coli.
- 5′ cap
- A structure at the 5′ end of mRNA. Needed to start translation, it also protects against degradation.
- UTR
- Untranslated region. Does not code for protein but strongly affects stability and translation efficiency.
- Poly(A) tail
- A run of adenines following the 3′ end of mRNA, involved in stability and translation.
- Pseudouridine
- An isomer of uridine. When incorporated, it makes RNA less readily detected by the immune system.
- N1-methylpseudouridine
- m1Ψ. Used in place of uridine in approved mRNA vaccines.
- dsRNA
- Double-stranded RNA. An aberrant IVT product and an impurity that acts to shut down translation.
- LNP
- Lipid nanoparticle. A dispersion of four lipids that encapsulates mRNA and carries it into cells.
- Ionisable lipid
- A lipid that is positively charged at low pH and neutral at physiological pH. The main component of an LNP.
- DSPC
- Distearoylphosphatidylcholine. Melting point about 54 °C; cylindrical, it forms a lamellar phase.
- PEG lipid
- A lipid bonded to polyethylene glycol. Affects particle size, aggregation and circulation time.
- Rapid mixing
- Making LNPs by bringing an ethanol phase and an aqueous phase together all at once, at a set pH and flow rate.
- Microfluidic mixing
- The leading form of rapid mixing, in which two liquids meet in very fine channels.
- Encapsulation
- The share of the added mRNA that ends up inside the particles. A release specification item.
- Polydispersity
- A measure of the spread in particle size, measured by dynamic light scattering.
- Cryoprotectant
- An additive that prevents damage from freezing and thawing, such as sucrose or trehalose.
13. References (primary sources)
- The Nobel Assembly at Karolinska Institutet "The Nobel Prize in Physiology or Medicine 2023 — Press release", 2 October 2023 — nobelprize.org
- Karikó, K., Buckstein, M., Ni, H., Weissman, D. "Suppression of RNA recognition by Toll-like receptors: the impact of nucleoside modification and the evolutionary origin of RNA", Immunity 23(2), 165–175 (2005). doi:10.1016/j.immuni.2005.06.008 — pubmed.ncbi.nlm.nih.gov
- Pardi, N., Hogan, M.J., Porter, F.W., Weissman, D. "mRNA vaccines — a new era in vaccinology", Nature Reviews Drug Discovery 17(4), 261–279 (2018). doi:10.1038/nrd.2017.243 — pmc.ncbi.nlm.nih.gov
- Hou, X., Zaks, T., Langer, R., Dong, Y. "Lipid nanoparticles for mRNA delivery", Nature Reviews Materials 6(12), 1078–1094 (2021). doi:10.1038/s41578-021-00358-0 — pmc.ncbi.nlm.nih.gov
- FDA "COMIRNATY (COVID-19 Vaccine, mRNA)" prescribing information (revised August 2026) — fda.gov
- European Medicines Agency (CHMP) "Comirnaty — Assessment report", EMA/707383/2020, 19 February 2021 — ema.europa.eu
- FDA "SPIKEVAX (COVID-19 Vaccine, mRNA)" prescribing information (revised August 2026) — fda.gov
- Leung, A.K.K., Tam, Y.Y.C., Chen, S., Hafez, I.M., Cullis, P.R. "Microfluidic Mixing: A General Method for Encapsulating Macromolecules in Lipid Nanoparticle Systems", The Journal of Physical Chemistry B 119(28), 8698–8706 (2015). doi:10.1021/acs.jpcb.5b02891 — pubmed.ncbi.nlm.nih.gov
- Andries, O. et al. "N(1)-methylpseudouridine-incorporated mRNA outperforms pseudouridine-incorporated mRNA by providing enhanced protein expression and reduced immunogenicity in mammalian cell lines and mice", Journal of Controlled Release 217, 337–344 (2015). doi:10.1016/j.jconrel.2015.08.051 — pubmed.ncbi.nlm.nih.gov
- FDA "COMIRNATY" product page (STN and list of approval letters) — fda.gov
- FDA "SPIKEVAX" product page (STN and list of approval letters) — fda.gov
14. Claim-to-source audit
| Claim in the text | Basis | Label |
|---|---|---|
| That the 2023 Nobel Prize in Physiology or Medicine was awarded jointly to Katalin Karikó and Drew Weissman for their discoveries concerning nucleoside base modifications, and that the announcement was made on 2 October 2023; that dendritic cells recognise in vitro transcribed mRNA as foreign, leading to activation and release of inflammatory signalling molecules; that including base modifications almost abolished the inflammatory response; that the work was published in 2005; and that two vaccines were approved in December 2020 with protective efficacy of around 95% reported | Nobel Prize press release (2 October 2023)[Reference 1] https://www.nobelprize.org/prizes/medicine/2023/press-release/ | Sourced |
| That RNA signals through human TLR3, TLR7 and TLR8; that incorporating m5C, m6A, m5U, s2U or pseudouridine abolishes that activity; the conclusion that nucleoside modification suppresses the ability of RNA to activate dendritic cells; and the journal, volume, issue and year | Karikó et al. (2005), Immunity, abstract[Reference 2] https://pubmed.ncbi.nlm.nih.gov/16111635/ | Sourced |
| That mRNA is a non-infectious, non-integrating platform with no potential risk of infection or insertional mutagenesis; that IVT mRNA is produced from a linear DNA template using T7, T3 or SP6 phage RNA polymerase; that the 5′ cap is added during or after transcription by the vaccinia virus capping enzyme or by synthetic or anti-reverse cap analogues; that the poly(A) tail is added directly from the DNA template or by poly(A) polymerase; that rare codons are replaced with frequently used ones and that sequence optimisation can raise G:C content; that UTRs are taken from viral and eukaryotic sequences and greatly increase half-life and expression; that enzymatically synthesised mRNA preparations contain dsRNA as an aberrant IVT product, whose recognition leads to type I interferon production and, via PKR and OAS, to translational inhibition and degradation of cellular RNA; that it can be removed by FPLC or HPLC; and that FPLC purification can raise protein production in primary human dendritic cells by up to 1,000-fold | Pardi et al. (2018)[Reference 3] https://pmc.ncbi.nlm.nih.gov/articles/PMC5906799/ | Sourced |
| That mRNA contains five elements (5′ cap, 3′ poly(A) tail, coding sequence, and 5′ and 3′ UTRs) that are critical to initiation, translation, termination, post-transcriptional modification and degradation; that cap analogues allow eIF4E-mediated initiation and can resist decapping enzymes; that poly(A) interacts with poly(A)-binding protein. The GMP manufacturing sequence (linearised DNA template, IVT, DNase digestion, capping, purification, then storage buffer, sterile filtration and freezing). That LNP–mRNA is produced by rapid mixing and shows a stable nanostructure, with mRNA encapsulated in the core by electrostatic interactions and protected from nuclease degradation. That ionisable lipids are protonated at low pH and neutral at physiological pH, that neutrality reduces interaction with blood-cell membranes, and that protonation in endosomes can promote membrane destabilisation and endosomal escape. That DSPC has a melting point of about 54 °C and a cylindrical geometry that forms a lamellar phase and stabilises particle structure, and that DOPE has a melting point of about 30 °C and a conical shape adopting the inverted hexagonal H(II) phase. That cholesterol can improve particle stability by modulating membrane integrity and rigidity. That the amount of PEG lipid affects particle size and zeta potential, reduces aggregation and can extend circulation time. That DLin-MC3-DMA is a key delivery component of Onpattro; the design approach of introducing ester bonds to increase biodegradability; that SM-102 and ALC-0315 show better in vivo delivery efficiency and pharmacokinetics than MC3; and that SM-102 and ALC-0315 are the ionisable components of mRNA-1273 and BNT162b respectively. The older methods (thin-film hydration, reverse-phase evaporation, extrusion) and current rapid mixing (ethanol and aqueous phases mixed at set pH and flow rate). That storage form and cryoprotectant type affect long-term stability; that adding 5% sucrose or trehalose maintained in vivo delivery efficiency for at least three months in liquid nitrogen storage; that the approved COVID-19 mRNA vaccines are all stored frozen in the presence of sucrose; and the outlook that formulations not needing cold or frozen storage would lower costs and speed up wider access | Hou et al. (2021)[Reference 4] https://pmc.ncbi.nlm.nih.gov/articles/PMC8353930/ | Sourced |
| The composition of COMIRNATY (2026-2027 Formula) per 0.3 mL: modRNA 30 µg, ALC-0315 0.43 mg, ALC-0159 0.05 mg, DSPC 0.09 mg, cholesterol 0.19 mg, tromethamine 0.06 mg, tromethamine hydrochloride 0.4 mg, sucrose 31 mg. The composition of the presentation for ages 5 to 11 (modRNA 10 µg, ALC-0315 0.14 mg, ALC-0159 0.02 mg, DSPC 0.03 mg, cholesterol 0.06 mg). That the active ingredient is described as nucleoside-modified messenger RNA (modRNA) and that the product is a sterile suspension for intramuscular use. That single-dose vials can be stored in an ultra-low-temperature freezer at −90 °C to −60 °C and must not be stored at −25 °C to −15 °C; that once moved to 2 to 8 °C they can be kept for up to 10 weeks and must not be refrozen; and that total time at 8 to 25 °C must not exceed 12 hours. That prefilled syringes must not be frozen. That the manufacturer is BioNTech Manufacturing GmbH | FDA COMIRNATY prescribing information[Reference 5] https://www.fda.gov/media/151707/download?attachment | Sourced |
| That the active substance is a single-stranded, 5′-capped mRNA, containing no uridine, with N1-methylpseudouridine used instead. That active-substance manufacture consists of five main steps, with IVT from linear DNA followed by purification and filtration steps, and a final filtration before dispensing and frozen storage. That the linear DNA template is produced via plasmid DNA obtained from transformed E. coli cells and, though not part of the final product, defines the sequence. The main drug product steps (thawing and dilution of the active substance, LNP formation and stabilisation, buffer exchange, concentration and filtration, concentration adjustment and addition of cryoprotectant, sterile filtration, aseptic filling, visual inspection, labelling, freezing and storage). That ALC-0315 and ALC-0159 are classed as novel excipients, that DSPC and cholesterol are used in approved products, and that DSPC is part of the LNP in Onpattro, which is given intravenously at higher doses. The active-substance specification items (RNA integrity by capillary gel electrophoresis, 5′ cap by RP-HPLC, poly(A) tail by ddPCR, residual DNA template by qPCR, dsRNA by immunoblot and others). The finished-product specification items (LNP size and polydispersity by DLS, RNA encapsulation and RNA content by fluorescence assay, lipid content and identity by HPLC-CAD, RNA integrity by capillary gel electrophoresis, potency / in vitro expression by cell-based flow cytometry and others). That the acceptance criteria for LNP size, polydispersity, RNA encapsulation, in vitro expression and RNA integrity were tightened during the procedure. That the transport temperature range of −90 to −60 °C is based on stability data, and that one thaw and refreeze cycle is permitted during transport | EMA public assessment report (Comirnaty, EMA/707383/2020)[Reference 6] https://www.ema.europa.eu/en/documents/assessment-report/comirnaty-epar-public-assessment-report_en.pdf | Sourced |
| The composition of SPIKEVAX (2026-2027 Formula) per 0.5 mL: nucleoside-modified mRNA 50 µg, total lipids 1.01 mg (SM-102, PEG2000-DMG, cholesterol, DSPC), tromethamine 0.25 mg, tromethamine hydrochloride 1.2 mg, acetic acid 0.021 mg, sodium acetate trihydrate 0.10 mg, sucrose 43.5 mg. That it is stored frozen at −50 °C to −15 °C and, after thawing, can be kept at 2 to 8 °C for up to 60 days. That the manufacturer is Moderna Tx, Inc. | FDA SPIKEVAX prescribing information[Reference 7] https://www.fda.gov/media/155675/download?attachment | Sourced |
| That LNPs composed of an ionisable cationic lipid, PEG lipid, DSPC, cholesterol and siRNA can be made efficiently by microfluidic mixing; that the method extends to encapsulating mRNA (1.7 kb) and plasmid DNA (6 kb); that raising the proportion of DSPC gives LNP–siRNA systems a more bilayer-like structure; and that the core can adopt a continuum of nanostructures depending on the proportions and structural preferences of the constituent lipids | Leung et al. (2015), abstract[Reference 8] https://pubmed.ncbi.nlm.nih.gov/26087393/ | Sourced |
| That m1Ψ-modified mRNA, alone or combined with m5C, outperformed Ψ and/or m5C/Ψ-modified mRNA, giving reporter gene expression up to about 44 times higher in cell lines (double-modified versus double-modified) and about 13 times higher in mice (single-modified versus single-modified) | Andries et al. (2015), abstract[Reference 9] https://pubmed.ncbi.nlm.nih.gov/26342664/ | Sourced |
| That the STN for COMIRNATY is 125742, its proper name is "COVID-19 Vaccine, mRNA", its manufacturer is BioNTech Manufacturing GmbH, and the earliest FDA approval letter is dated 23 August 2021 | FDA COMIRNATY product page[Reference 10] https://www.fda.gov/vaccines-blood-biologics/comirnaty | Sourced |
| That the STN for SPIKEVAX is 125752, its proper name is "COVID-19 Vaccine, mRNA", its manufacturer is Moderna Tx, Inc., and the earliest FDA approval letter is dated 31 January 2022 | FDA SPIKEVAX product page[Reference 11] https://www.fda.gov/vaccines-blood-biologics/spikevax | Sourced |
| Putting total lipid in Comirnaty at 0.76 mg, lipid to mRNA by mass at about 25 times and sucrose to mRNA at about 1,000 times; putting the lipid mass composition at ALC-0315 about 57%, cholesterol 25%, DSPC about 12% and ALC-0159 about 7%; putting lipid to mRNA in Spikevax at about 20 times and sucrose to lipid at about 43 times; and the total lipid of 0.25 mg in the 10 µg presentation, a lipid-to-mRNA ratio of about 25 times, the same as the 30 µg presentation | Our calculation. These are simple sums and divisions of the values stated on the label, not the input molar ratios used in manufacture, and they say nothing about efficacy or safety | Our calculation |
| Whether mRNA–LNP formulations not needing cold or frozen storage have been achieved | At the time of writing (September 2026) this article could not confirm any primary source showing that this has been achieved | Not yet confirmed |
| The mass of each of the four lipids in Spikevax | The FDA prescribing information gives only total lipids, so this article has not calculated per-lipid ratios | Commentary |
| LNP manufacturing conditions (specific pH and flow-rate values), molar ratios, equipment configuration and manufacturing cost | Not stated in this article because the primary sources it consulted do not publish specific values | Commentary |
| The indications, clinical results and safety assessment of each product | This article explains materials and manufacturing technology and does not assess the efficacy or safety of treatments or vaccines. For approved products it quotes only statements on composition, manufacture and handling that appear in material published by the regulators. The efficacy figure at the end of Section 11 is quoted as stated in the Nobel announcement | Commentary |
| Reading the lipid ratios as a functional-material formulation sheet; the division of roles into main ingredient, framework, stiffness control and surface modification; likening the relation between molecular shape and phase behaviour to the critical packing parameter; the reading that the same formulation mixed differently gives a different particle; likening the sensitivity to dsRNA removal to high-purity materials; reading from the list of specification items that these quantities can shift with process conditions; the reading that novel-excipient status affects development cost; and placing sucrose as the heaviest component | Our summary and commentary based on published content. Not views expressed by the authors of the papers or by the regulators | Commentary |
| That Figs. 1, 2, 3, 5 and 6 are explanatory drawings rather than real molecular structures, equipment or measurement images, and that the hero image and Fig. 4 are AI-generated images | Our note | Commentary |
Last updated 23 September 2026. Sources are limited to primary material (the Nobel Foundation announcement, FDA-approved prescribing information, the EMA public assessment report and peer-reviewed papers). Because the article includes readings from a formulation and process-design standpoint, those are marked as Commentary and kept separate from sourced fact. Specific LNP manufacturing conditions (pH, flow rate, molar ratios), equipment configuration, manufacturing cost and the lipid breakdown of Spikevax are not stated here because they could not be confirmed in published primary sources. This article explains materials and manufacturing technology; it does not assess the efficacy or safety of any treatment or vaccine, and it is not medical advice. All figures are explanatory concept graphics. Figs. 1, 2, 3, 5 and 6 are vector drawings; the hero image and Fig. 4 are AI-generated images, and none of them is a photograph of a real molecule, particle or product.