Eshmuno^® Fit Custom Affinity Resin Development Services: Frequently Asked Questions

Find answers to frequently asked questions below, or for more details see Eshmuno^® Fit Custom Affinity Resin Development Services.

How does the Eshmuno^® Fit custom affinity technology work?

The Eshmuno^® Fit custom affinity technology uses Nanofitin^® ligands, specifically designed to bind to a target molecule, such as a protein or other biological target. These custom ligands are immobilized onto the Eshmuno^® chromatography matrix, creating a custom affinity resin. This technology allows for the selective capture and purification of the target molecule from a complex mixture, such as a clarified cell lysate or cell culture supernatant.

What are Nanofitin^® ligands?

Nanofitin^® ligands are small, single-chain, cysteine-free affinity proteins derived from the Sac7d protein, isolated from the hyperthermophile archaeon discovered in the Yellowstone USA National Park geysers. Nanofitin^® proteins frame a versatile platform for custom-generating ligands for affinity separation. They fold into a well-characterized OB-fold and share stability profiles that make them suitable for various applications, including as custom affinity ligands for purification processes.

How do Nanofitin^® library generation and screening work?

The Nanofitin^® ligand library is screened through a process called ribosome display, which involves the in vitro selection of high-affinity ligands from a combinatorial library of Nanofitin^®. The library is generated by randomizing the natural DNA-binding site of the Sac7d protein, yielding a diverse set of Nanofitin^® with unique binding properties. These Nanofitin^® ligands are then screened for their ability to bind to a specific target molecule through multiple rounds of selection, and the best candidates are further characterized for their expression yield, binding kinetics, and stability under cleaning-in-place (CIP)-like conditions.

Can the Eshmuno^® Fit resin be customized for a specific molecule or a cluster of molecules?

Yes, Eshmuno^® Fit resin can be customized for a specific molecule or cluster of molecules. This process typically includes ligand generation and selection, Nanofitin^® ligand expression and purification, and evaluation of the custom affinity resin to ensure it meets the required specifications for downstream applications.

How is the optimal ligand determined for the Eshmuno^® Fit custom affinity resin?

The optimal ligand for Eshmuno^® Fit custom affinity resin is determined through a process of ligand generation and selection, ribosome display selection, enzyme-linked immunosorbent assay (ELISA) screening, sequencing, expression, purification, and validation of binding kinetics and stability under cleaning-in-place (CIP)-like conditions. This comprehensive process ensures that the optimal ligand is chosen for the Eshmuno^® Fit custom affinity resin, providing high binding capacity and tailored stability in caustic solutions.

What is the expected binding capacity and resolution of the Eshmuno^® Fit resin?

The typical binding capacity is usually in the double-digit milligrams per milliliter range, and the resolution is influenced by the ligand to target specific affinity constants of < 10^-7 M. However, this varies depending on the particular ligand and target molecule used in the purification process. 

Can the resin be scaled up for larger manufacturing processes?

Yes, the scalability of the Eshmuno^® Fit resin was investigated for larger manufacturing processes. A template process has been developed to flexibly scale custom affinity resins from the research scale to the final lot size required for industrial applications.

What quality control criteria are in place for the custom affinity resins?

The custom affinity resins’ quality control criteria include the following parameters: appearance, microscopic evaluation, particle size, dynamic binding capacity (DBC), bicinchoninic acid (BCA) protein assay, dynamic binding capacity, total yeast and mold count (TYMC), total aerobic microbial count (TAMC), and endotoxins.

Are Regulatory Support Files (RSF) available for Eshmuno^® Fit?

Regulatory Support Files are available for Eshmuno^® Fit custom affinity resins once the project has proceeded to  full-scale manufacturing.

Have toxicity studies of the ligand been performed?

Certainly! We conducted two toxicity studies. The first study involved 8 mg/kg/day and 80 mg/kg/day dosages for 5 days, while the second study included an additional 400 mg/kg/day dosages for 14 days. We used an artificially created Nanofitin^® ligand with a high immunogenicity score for the study. The results will be published soon and, in the meantime, are available upon direct request.

How can the leached ligand be detected?

The leached Nanofitin^® ligand can be detected using the ELISA assay, which encompasses two different assays applicable to different detection ranges. Alternatively, high-resolution liquid chromatography coupled with mass spectrometry can be applied to detect leached Nanofitin^® ligands.

How does the cost of custom affinity chromatography resins compare to standard off-the-shelf resins?

While custom affinity chromatography resins like Eshmuno^® Fit may initially pose a higher cost than standard chromatography resins, their customized nature offers unparalleled advantages. The long-term benefits of custom affinity resins, such as streamlined downstream processing, improved yields, and reduced cost of goods, can lead to significant cost savings over time. Implementing Eshmuno^® Fit is not just about immediate costs but about enabling templated purification, securing enduring benefits, and optimizing your operations for sustained success.

Is there a value model that provides expected costs for evaluating the economic feasibility of working with this solution?

Yes. The value model for Eshmuno^® Fit custom affinity resin development services is designed to quantify the benefits and long-term value creation of Eshmuno^® Fit for potential adopters. The model aims to support informed decision-making by comprehensively analyzing the qualitative and quantified benefits and comparing them with traditional processes. The model includes factors such as time to market, process simplification, yield, cost savings, sustainability, and other measurable benefits. It provides scenarios for long-term value creation, cost savings, payback periods, and Net Present Value (NPV) for specific projects.

What is the turnaround time for producing and delivering custom affinity resins?

The turnaround time for producing and delivering custom affinity resins involves several stages and can take several months, depending on the specific requirements and the need for additional rounds of screening or optimization. A flexible project structure makes it possible to achieve clearly defined deliverables after each project stage e.g,. small scale prototypes, research scale samples or analytical test results.

Does our company assist with process optimization, integration, and addressing challenges that may arise during implementation?

Eshmuno^® Fit is a service that is offered in the form of a customized project. Based on the pre-developed underlying technology, the project includes continuous dialogue with the client as well as the opportunity to address specific requirements. The aim is to develop a bespoke product that is optimally adapted to the specific requirements of the customer's process. Our application specialists are available to provide additional support during the implementation of the product.

Is there a business continuity plan in place for the custom affinity resins?

Yes, the Eshmuno^® Fit manufacturing site has a business continuity plan in place to ensure it continues operating during and after unexpected events or disruptions. 

Is there a minimum order quantity for Eshmuno^® Fit custom affinity chromatography resins?

As every Eshmuno^® Fit product is customized, the purchased quantity can be adapted to customer requirements.

What are the intellectual property considerations for custom affinity resins?

Since our company has exclusive rights to Nanofitin^® ligand technology in the downstream field, this enables us and the customer to look for a common agreement for the intellectual property questions. 

How much material feed is required to develop a Nanofitin^® ligand?

The Nanofitin^® ligand selection process is downscaled to such a level that only several milligrams of purified target are required. Depending on the project scope, additional material might be required, especially for target molecules in the presence of process and product-related impurities, to enable a selection of the best Nanofitin^® ligand. The standard project scheme assumes that customers test small-scale prototypes.  

What is the process for qualifying the technical and financial feasibility of a Nanofitin^® project?

The initial step involves filling out the inquiry form on the Eshmuno^® Fit webpage or contacting your sales representative. Subsequently, your sales representative will coordinate with the Eshmuno^® Fit team, which includes research and development (R&D), product management, and project management, to introduce the Eshmuno^® Fit custom affinity development services program. This involves gathering information on your requirements, addressing your queries, and assessing the program's feasibility. Additionally, value managers may participate in these discussions to create an economic evaluation model for the project, and legal partners may be involved in drafting the service agreement. This support extends throughout the implementation phase.

Can Nanofitin^® ligands be coupled to Natrix^® membranes?

No. Currently, there is no commercial platform for producing custom affinity membranes.  

Nanofitin^® is a registered trademark of Affilogic.