Product – E&S Adjuvants

Our Product

ES-AdjuVac

We help you overcome the challenges of modern veterinary vaccines development

Compared to Incomplete Freund Adjuvant (IFA)
Recommended for water in oil adjuvant used for low viscosity vaccines formulations.
This adjuvant produces a low viscosity emulsion.
Can be used with formulations from 50 – 70% ratio water/oil.
Compatible with Saponin.
Suitable for Bovines, Poultry, Small ruminants, and Swine

Emulsion preparation protocol

I. Preparation oil phase

- Weigh 1% of the final emulsion volume of AdjuVac.
- Weigh the Mineral oil to the correspondent oil phase
- Mix both components under agitation.
- Filter the oil phase by a 0.22-micrometer filter into a sterile vessel or heat sterilize at 121 °C for 1 hour.

II – Preparation water phase

Weigh the required amount of water phase with the antigen and other components.

III. Preparation emulsion

Put the oil phase under agitation using the colloid mill (refrigerated 2-8°C)
Add the water phase (refrigerated) to the oil phase while emulsifying (using the colloid mill).

Note: Mixture the water phase slowly but constantly

The emulsification process is continued until all the water phase is mixed
Keep the colloid emulsifying for an extra 3 to 5 minutes to ensure complete emulsification
If homogeneous, cool the emulsion under stirring as quickly as possible to a temperature of 2 – 8°C.
Store the final bulk at 2-8°C.

Exemple – Preparation 1 Liter of AdjuVac:

Preparation 1000g
Oil Phase	Mineral Oil	500
	AdjuVac	10
Water Phase		490
Total		1000

AdjuVac Preparation

How to evaluate the emulsion:

A total of 4 tests can be performed in the after the emulsification process:

Droplet test: The droplet dispersal test is performed for the vaccine to check whether a proper emulsion was obtained. Simple pipette a droplet of vaccine and put it in a beaker with water; if the oil droplet is stable, that means the process was performed correctly
Centrifugation test: Centrifuge the vaccine at 3000 rpm for 30 minutes and check for phase separation. The phases cannot exceed 5% separation
Long term stability: Put a sample of the product at 37°C and 4°C and check daily for phase separation
Microscope evaluation: Put 1 droplet in a microscope and evaluate the emulsion characteristics as below:

Emulsion evaluation under microscope 100 x magnitude

Testing of the AdjuVac - Field experiments

Foot-and-mouth disease (FMD) is a highly contagious and rapidly spreading disease of cloven-hoofed animals. Animals are immunized with inactivated whole virus vaccines to control the spread of foot-and-mouth disease virus (FMDV). In Brazil the Government campaigns together with the industry efforts have been shown to be very efficient in the prevention of FMD, therefore we design the study to compare the ES-AdjuVac, with the commercial available adjuvants.

Material and methods

Total of 3 groups of six FMD-free cattle were vaccinated with FMD vaccine formulations 1–3 (Table 1). Animals vaccinated with vaccine formulation 1-3 were vaccinated, samples were collected at 4-week intervals for serological testing and local reaction measurements performed daily.

Table 1

Adjuvant	Imunostimulant	Vaccine Antigens	Application	Dose volume
ES-AdjuVac with saponin	Saponin 1 mg/dose	Virus A24 Cruzeiro, O1 Campos, C3 Indaial	Subcutaneously	5 ml
Competitor	Saponin 1 mg/dose	Virus A24 Cruzeiro, O1 Campos, C3 Indaial	Subcutaneously	5 ml
ES-AdjuVac without saponin	No saponin	Virus A24 Cruzeiro, O1 Campos, C3 Indaial	Subcutaneously	5 ml

Table 2

At 28 days after vaccination animals were tested for EPP – expectancy of protection (ELISA), with threshold of 80%. (Considering >80%) protected animals.

Formulation	Sorotype O	Sorotype A	Sorotype C
ES-AdjuVac with saponin	96,66	92,38	82,13
Competitor	90,32	80,53	82,89
ES-AdjuVac without saponin	93,32	80,53	89,72

Table 3

At 56 days after vaccination animals were tested for non-structural proteins after the re-vaccination.

Non-structural proteins - 28 days (post revaccination)
	Tests
Formulations	ELISA 3 ABC	EITB
ES-AdjuVac with saponin	0/18	Non-Reagent
Competitor	0/18	Non-Reagent
ES-AdjuVac without saponin	0/18	Non-Reagent

Graphic 1

At 56 days after vaccination animals were tested for non-structural proteins after the re-vaccination.

Results and Conclusion

Our ES-AdjuVac is highly effective against all viruses (as tested using in-house ELISA) with an efficacy rate above the 80% threshold. Moreover, it does not produce any reactivity to non-structural proteins (as assessed by the in-house assay developed by Panamerican PANAFTOSA). At 28 days after vaccination, the average local reaction in animals was 10 cm², compared to the competitor’s 50 cm². This data demonstrates that our adjuvant can produce a robust immune response while minimizing local reactions. It is crucial to minimize such reactions as they can compromise weight gain, milk production, growth, and in some cases, even lead to fatal consequences. Our adjuvant is safe for bovines, even with saponin use, and produces a solid immune response while reducing local reactions.

How are we different?

Our products are safe, reliable, and proven efficacious in the field, and our company will be present in all the process steps, from development to scale-up and the entire life cycle.