Evidence reviewed through: July 2026
What this page examines. Cell count is the number most clinics lead with, and it is the easiest
to compare. This page sets out what the published evidence actually establishes about the two
variables people conflate — how many cells and what state those cells are in — and is
explicit about where the evidence is strong, where it is contested, and where it does not exist.
Product specifications are stated as what they are: commitments about what we supply, not research
findings.
Key Points
- Viability and metabolic fitness are the dominant quality attributes. This is the ISCT MSC committee's own position, not a marketing claim.
- Cells administered immediately after thawing are functionally impaired. In a mouse colitis model, viable cells given straight from the thaw performed like heat-inactivated cells.
- That impairment is largely reversible. Twenty-four hours of post-thaw culture fully restored immunosuppressive function in the study that identified the defect.
- Some cryoinjury is not reversible. Thawed cells show a homing and biodistribution defect that licensing strategies do not fully rescue.
- But optimised cryopreservation can match fresh. Several studies find no meaningful difference when post-thaw viability is preserved — this is a genuinely contested question, not a settled one.
- More cells is not better. The best-controlled dose-ranging trial in knee osteoarthritis found the lowest dose most effective, with adverse events concentrated in the higher-dose groups.
- No trial has ever compared 50 million fresh cells against 100 million cryopreserved cells. Dose and preservation state are separate variables, and the direct comparison has not been tested.
1. The Question, Stated Precisely
"100 million cells" and "50 million cells" are easy to compare. That is exactly why the number is advertised — and why it is the wrong place to stop.
Two independent variables sit behind any cell product:
- How many cells are administered.
- What functional state those cells are in when they reach the patient.
These are frequently blurred together. They should not be, because the evidence on each points in a different direction, and no study has tested them in combination. There is no trial comparing 50 million fresh cells with 100 million cryopreserved cells, at any indication. Anyone claiming a settled answer to that specific comparison — including anyone selling either configuration — is going beyond the evidence.
What the literature does support is more interesting than the headline number.
2. Viability and Fitness Are the Dominant Quality Attributes
This is not a marketing position. It is the stated view of the International Society for Cell & Gene Therapy's MSC committee, which proposes that "MSC viability and metabolic fitness likely dominate as a potency quality attribute," and that in-vivo persistence affects potency in ways that in-vitro surrogate assays do not predict. [1]
The experimental basis is striking. In a mouse model of toxic colitis:
- Replication-fit MSCs harvested in log phase of growth produced substantial improvement in clinical and pathological endpoints.
- Heat-inactivated (dead) MSCs lost all therapeutic utility — and "the observation is mirrored by use of viable MSC administered immediately post-thaw from cryostorage."
- Transient in-vivo engraftment was "severely compromised when MSCs are dead or thawed."
- The maximum tolerated intravenous bolus dose failed to produce the benefit that a smaller, well-delivered dose achieved.
The authors' conclusion: "pharmaceutical potency of MSC requires viability and functional fitness," and intravenous administration of thawed products "may be biased against positive clinical outcomes." [2]
That last point deserves emphasis. In that model, the highest tolerated dose did not rescue a delivery problem — and viable-but-just-thawed cells behaved like dead ones.
3. The Thaw Problem — and What Fixes It
3.1 What happens at thaw
The mechanism is well characterised. Freshly thawed cryopreserved MSCs:
- up-regulate heat-shock proteins;
- become refractory to interferon-γ-induced IDO up-regulation — the pathway through which MSCs suppress T cells;
- are compromised in suppressing T-cell proliferation. [3]
Separately, thawed MSCs are lysed on coculture with activated peripheral blood mononuclear cells, an effect enhanced with allogeneic cells. DMSO-free cryoprotectants, substituting human platelet lysate for human serum albumin, and autophagy or caspase inhibitors did not prevent these thaw defects. [4]
Freeze-thawed cells also trigger the instant blood-mediated inflammatory reaction more strongly and activate complement, leading to faster clearance after infusion. [5]
3.2 What fixes it — and what does not
Most of the functional defect is reversible. In the study that identified it, immune suppressor activity, interferon-γ responsiveness and IDO induction were "fully restored following 24 h of MSC tissue culture post-thaw." The authors framed this as a probable explanation for the underperformance of MSC therapy in trials using cryopreserved cells thawed immediately prior to infusion. [3]
Some of it is not. Interferon-γ prelicensing before cryobanking restores T-cell suppression to the level of fresh cells — but prelicensed thawed MSCs "do not deploy lung tropism in vivo following intravenous injection as well as fresh MSCs." The authors identify both reversible and irreversible cryoinjury mechanisms, affecting cell survival and tissue distribution. [4]
So the honest formulation is not "frozen cells are damaged." It is: cells given straight out of the thaw are functionally impaired; recovery culture restores most but not all of that function; and biodistribution appears to be the part that does not fully recover.
4. Where Cryopreservation Is Not the Problem
Applying the same scrutiny in the other direction, there is a substantial body of evidence that optimised cryopreservation does not meaningfully degrade cells.
- Using modified cryopreservation methods, MSCs retained >95% post-thaw viability, remained responsive to inflammatory signals, suppressed activated peripheral blood mononuclear cells, and "performed as well as fresh MSC" in an in-vivo ischaemia/reperfusion model. [6]
- A GMP-grade comparison concluded that cryopreservation does not alter the main characteristics of clinical-grade MSCs, including immunomodulating potential. [7]
- A systematic review of 18 comparative preclinical studies — 257 in-vivo efficacy experiments across 101 outcome measures — found only 2.3% significantly different, and of those six, two favoured fresh and four favoured cryopreserved. [8]
A review posing the question directly is titled "Cryopreserved or Fresh Mesenchymal Stromal Cells: Only a Matter of Taste or Key to Unleash the Full Clinical Potential of MSC Therapy?" — which is a fair summary of where the field sits. [9]
This is a genuinely contested question. It would be as wrong to conclude "fresh is categorically superior" as to conclude "preservation state does not matter." What reconciles the two bodies of evidence is how the product is handled: the pro-fresh findings concentrate on cells administered immediately post-thaw, and the null findings concentrate on optimised protocols where post-thaw viability is preserved.
5. What Actually Matters: Readiness at the Point of Administration
The useful distinction is not fresh versus frozen as categories. It is whether the cells are functionally ready when they enter the patient.
The failure mode worth avoiding is the third row — and it is common, because it is operationally convenient. Preparing cells fresh for same-day administration sidesteps the question entirely. So does cryopreservation done properly with recovery culture. What does not work is thawing a vial at the bedside and injecting it.
This is why the question to ask is not "fresh or frozen?" but "what was the viability at the moment of administration, and how long between thaw and injection?"
6. On Cell Count: More Is Not Better
The dose evidence does not support a simple "more cells" rule — in either direction.
- In a randomised placebo-controlled dose-ranging trial in knee osteoarthritis, a trend toward improvement was seen in the 25-million-cell group across VAS, ICOAP and WOMAC — not statistically significant versus placebo — while "adverse events were predominant in the higher dose groups (50, 75, and 150 million cells)." Whole-organ MRI showed no difference from baseline or placebo. The authors concluded: "A twenty-five-million-cell dose may be the most effective among the doses tested." [10]
- An analysis of 914 registered MSC trials found minimum effective intravenous doses clustering around 100–150 million cells, *"with lower and higher doses being less effective."* [11]
- A network meta-analysis of 11 randomised knee osteoarthritis trials found moderate doses produced the greatest long-term improvement, with high doses producing more adverse events. [12]
- In the colitis model above, the maximum tolerated intravenous bolus failed where a smaller, better-delivered dose succeeded. [2]
What this supports: dose sits in a window rather than on a ladder, and higher counts carry a measurable adverse-event cost without a corresponding benefit.
What this does not support: that 50 million cells outperform 100 million cells. No study has tested that comparison, and the dose literature points toward an optimum range rather than a ranking. The defensible statement is that a well-characterised 50-million-cell dose sits within the range where trials have found benefit, and that higher counts are not evidence of a better product.
7. Donor Age
Donor age genuinely affects MSC biology. Age-related decline in proliferation, differentiation capacity and antioxidant defence in human bone-marrow MSCs is well documented, [13][14]and Wharton's jelly–derived cells have greater expansion capability and faster growth in vitro than bone marrow MSCs. [15]
Two honest qualifications belong alongside that:
- The authors of the Wharton's jelly review note that "further work is needed to determine whether WJCs engraft long-term and display self-renewal and multipotency in vivo." [15]
- At the level of clinical outcomes, no definitive advantage of autologous over allogeneic MSCs has been demonstrated — a statement that cuts against superiority claims in either direction. [16]
So neonatal cells have measurable laboratory advantages in expansion and growth. Whether that translates into better patient outcomes has not been established by head-to-head clinical comparison.
8. What We Specify
The following are product specifications — commitments about what is supplied. They are business facts, not research findings, and no citation is attached to them because none would be appropriate:
The Certificate of Analysis records cell count, viability with testing method and date, immunophenotype, sterility, endotoxin and mycoplasma results, and processing date and batch number.
Why we consider this configuration defensible: it avoids the failure mode the evidence identifies most clearly — cells administered without functional readiness — and it sits within the dose range where trials have found benefit rather than above it. That is a reasoned position about process. It is not a claim that this configuration has been shown superior in a clinical trial, because no such trial exists.
9. Questions to Ask Any Clinic
Including us.
On viability
- What is the measured viability at the point of administration, not at release?
- What method was used, and on what date?
On preservation and timing
- Fresh or cryopreserved?
- If cryopreserved: what was post-thaw viability, and how long between thaw and injection? This is the question the evidence most clearly supports asking.
- Was recovery culture performed after thawing?
On dose and source
- What is the guaranteed cell count — not the "up to" figure?
- What tissue source, and what passage number?
On documentation
- Can I see the Certificate of Analysis for my specific batch?
A provider who answers the thaw-to-injection question specifically is engaging with the variable that has the clearest evidence behind it.
10. What This Page Does Not Claim
Stated plainly, because the distinction matters:
- That fresh cells are categorically superior to cryopreserved cells. Optimised cryopreservation can match fresh on most measured outcomes.
- That 50 million cells have been shown to outperform 100 million cells. No trial has compared them.
- That any specific viability figure translates into a proportional difference in clinical outcome. The relationship between viability percentage and patient benefit has not been quantified, and any formula purporting to convert one into the other is a model, not a measurement.
- That competitors deliver lower-viability product. We have no verified data on other providers' release specifications and do not assert any.
The evidence supports a narrower and more useful claim: functional readiness at the point of administration is the quality attribute that matters most, and it is the one patients are least often given data about.