Monoclonal antibodies remain a widely employed biotherapeutic. Due to their high affinity, operability, scalability, and regulatory track record, Protein A fillers are the initial step in downstream purification for nearly all commercially approved mAb manufacturing processes. Seebio continues to enhance and innovate its Protein A filler products to better cater to customer demands.

 

 

Protein A

Protein A (SPA), a membrane protein from Staphylococcus aureus, possesses the unique ability to bind specifically to antibodies. Protein A affinity chromatography columns have become a widely adopted method for purifying antibodies. These columns effectively separate and purify antibodies of various mammalian subtypes or genetically engineered recombinant proteins containing antibody Fc fragments from ascites, serum, cell culture supernatant, or cell extracts.

 

Natural Protein A boasts a stable structure, is not acid- or alkali-resistant, lacks disulfide bonds within its molecule, has five highly homologous regions, and exhibits binding effects with IgG. Protein A demonstrates a robust affinity for IgG from diverse sources, with the binding site located in the Fc segment of IgG, leaving its antigen-binding activity unaffected. Currently, affinity chromatography media utilizing Protein A as an affinity ligand are extensively used in the purification of monoclonal antibodies and Fc recombinant fusion proteins.

 

Figure 1: Staphylococcus aureus Protein A

 

The Protein A filler serves as the pivotal step in antibody purification. To achieve reusability and cost savings on chromatography media, it is essential to perform clean-in-place (CIP) during the purification process. While sodium hydroxide (0.1-0.5 M) solution is highly effective as a cleanser, natural Protein A cannot withstand high alkaline concentrations due to its sequence limitations, often resulting in denaturation and shedding.

 

Through genetic engineering, Seebio Biotech has eliminated the IgG non-binding region of natural Protein A, leaving only the Fc-binding domain. Furthermore, mutations and transformations have been introduced, along with an anti-protease linker, to minimize steric hindrance and enhance the functionality of rProteinA. This has significantly improved its acid and alkali resistance while maintaining a high dynamic binding capacity. Purified through chromatography, this recombinant protein is used in affinity chromatography columns, ensuring the purity of the final product by eliminating the potential adulteration of irrelevant IgG.

 

 

Introduction to Seebio® Recombinant Protein A (rProtein A)

 

Product	Seebio® Recombinant protein A (rProtein A）
Item number	ECL0019H
Purity（SDS-PAGE）	≥95%
Source	E.coli
Molecular weight	28.2kDa
Exterior	Lyophilized powder
Label	His tag

 

 

Properties and Uses of Seebio® Recombinant Protein A

 

The C-terminus of Seebio® rProtein A is equipped with cysteine, allowing for directional coupling to solid separation media, essential for the purification of monoclonal or polyclonal antibodies. This recombinant protein boasts the capability to bind to numerous human or mouse immunoglobulin subclasses (e.g., human IgG1, IgG2, IgG4; mouse IgG2, IgG2a, IgG2b, IgG3) as well as total immunoglobulins derived from cattle, hamsters, and rabbits.

 

Remarkably, Seebio® rProtein A can be coupled with a wide range of reporter molecules (including fluorescent dyes, enzyme labels, biotin, colloidal gold, and radioactive labels) without compromising its antibody-binding activity. These coupled protein A derivatives find applications in various antibody detection techniques such as Western blot, ELISA, and immunohistochemistry.

 

 

Advantages of Seebio® Recombinant Protein A

 

1 Non-antibody column purification ensures that the product remains uncontaminated by other antibodies, such as IgG.

2 It can withstand 0.5 M NaOH and 0.5 M HCl treatment without any degradation or loss of antibody binding capacity.

3 High pH elution significantly reduces IgG aggregation.

4 The product incorporates a linker group specifically designed for antibody purification columns, enabling direct use in the preparation of protein A columns.

5 Stable quality: Batch production ensures consistent and continuous production, eliminating any variation between batches and maintaining stable quality.

6 High purity: Minimal residual host protein.

7 Freeze-dried powder format: Easy to store and transport.

8 Compliance with Regulatory Requirements: The production equipment and environment adhere strictly to relevant regulatory standards, and the entire production process follows the ISO 9001 quality management system.

 

 

Product Description of Seebio® Recombinant Protein A

 

Stability:

The lyophilized powder of recombinant protein A remains stable for 24 months when stored at 2-8°C. Upon dissolution in PBS (pH 7.4), it is ready for coupling and should be used promptly. Once dissolved in water, it can be stored at or below -20°C.

 

 

Storage:

 

 

Product List:

 

Item number	Product	Packaging
ECL0019H	Seebio® Recombinant Protein A (rProtein A)-Specialized for chromatography packing	10mg
		100mg
		1g
		1kg

 

 

Attached table: Comparison of antibody binding abilities of Protein A, Protein G, and Protein L

 

Species	Subcategory	Protein A	Protein G	Protein L
people	IgG	+++	+++	+++
	IgG1	++++	++++	++++
	IgG2	++++	++++	++++
	IgG3	-	+++	+++
	IgG4	++++	++++	++++
	IgM	+	-	+++
mouse	IgG	++	++	+++
	IgGl	+	++++	+++
	IgG2a	++++	++++	+++
	IgG2b	+++	+++	+++
	IgG3	++	+++	+++
	IgM	-	-	+++
rabbit	IgG	+++	+++	+
goat	IgG	+	++	-
	IgG1	+	+++	-
	IgG2	+++	+++	-
sheep	IgG	+	++	-
	IgGl	+	++	-
	IgG2	+++	+++	-
rat	IgG	+	++	+++
	IgGl	-	+	+++
	IgG2a	-	++++	+++
	IgG2b	-	++	+++
	IgG2c	++	++	+++
	IgG3	+	++	N/A
dog		++	+	N/A
pig		+++	++	+++
kappa chain antibody		-	-	+++

 

Note: + means binding ability, the more +, the stronger the binding ability; - means no binding; N/A: No data