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Ab-Capcher ExTra™

Background

Ab-Capcher ExTra is an alkali-resistant affinity medium for IgG purification, which is coupled with an alkali-resistant Protein A-derivative (Protein A-R28) designed for higher IgG-binding capacity than that of Ab-Capcher. The dynamic binding capacity of Ab-Capcher ExTra is apporoximately 70 mg human IgG/mL of medium at 100 cm/min. Ab-Capcher ExTra shows higher binding to mouse and rat IgGs exceeding Ab-Capcher. Since Pyrogen testing is carried out on an every lot of manufacturing, antibodies purified with Ab-Capcher ExTra are suitable for cell-based assay*.

*Recommended that endotoxin-free buffers are used for purification of IgG with Ab-Capcher ExTra.

 

Protein A-R28 is an alkali-tolerant IgG-binding protein derived from protein A, which is developed with ProteNova’s patent technology. Protein A-R28 strongly binds to various species and subclasses of IgG, compared with Protein A and G. The coupling to resin (Ab-Capcher) provides an alkali-washable unique affinity medium with high binding capacity for immunoglobulin, which is useful for purification of human, rabbit, and mouse IgGs including mouse IgG1.Ab-Aapcher is also useful for immuno-precipitation experiments.

Binding Characteristics of Ab-Capcher (ExTra)
Species Subclass Ab-Cap Protein A Protein G
Mouse IgG1
IgG2a
++++
+++++
+
++++
++
+
Rat IgG1
IgG2a
++++
-* 
-
-

+
+

Goat IgGs ++++ - +
Chicken IgY - - -
Human IgG +++++ ++++ ++
Rabbit IgG +++++ ++++ ++

*Connection has not confirmed.

Feature and Advantages

◊Dynamic binding capacity (100 cm/hr) is 70 mg/mL.
◊Reusable by washing with alkali.

Comparison of binding capacities

Static binding capacity
(Maximum binding capacity)
Excess of human polyclonal IgG was applied to the gel, shaked for 1 hr at RT, washed and eluted at pH 2.8. Amount of IgG in the elute was measured.
Dynamic binding capacity
(Applied to a column)
Human polyclonal IgG (3 mg/mL) was applied to a column (5x100 mm) at linear velocity of 100 cm/hr (0.33mL/min). DBC at 10% brealthrough was determined. 
PTN_P0032_3

Application

Rat/Mouse/Human IgG purification

Specifications

Gel matrix : 6% highly closslinked agarose
Average particle size : 35 μm
Ligand : Alkali-resistant Protein A-derivative (Protein A-R28)
Ligand-coupling : secondary amine, multiple-point attachent
Binding capacity  
     Static : 100 mg Human IgG/mL of medium
    20 mg Mouse IgG1/mL of medium (purified from mouse ascitic fluid)
     Dynamic** : 85 mg Human IgG/mL of medium (60 cm/hr) 
  70 mg Human IgG/mL of medium (100 cm/hr) 
Maximum linear velocity : 500 cm/hr 
Pecommended liner velocity : 50-150 cm/hr 
Washing conditions : 0.1-0.5 M NaOH
Pyrogen testing : Endotoxin negative (Gel-Clot Technique) 
Preservative : 20% Ethanol 
Storage  : 4-8°C 

**Determined at 10% breakthrough on a column (5x108 mm, 2.1 mL) with 3 mg human IgG/mL

Application Data

1. Purification of mouse IgG from low-concentration samples by repeated addition

As simulation of purification from cultured medium, mouse monoclonal IgG1 at low concentration was purified by repeated addition of the sample. According to the protocol of Ab-Rapid SPiN, 0.5 mL of purified mouse IgG1 (0.1 mg/mL) was added to a SPiN column set in a centrifugal machine, stand for 4 min with sometimes mixing and centrifuged. These steps were repeated for 20 times. Total 10 mL of samples was added to the column, washed and eluted at pH2.8. Recovery (%) with Ab-Capcher ExTra us higher than that og Ab-Capcher, indicating that increasing dispersity of the medium at smaller particle size of 35 μm, influences the recovery of IgG.

Gel volume: 100 μL/column
Sample: Purified mouse IgG1 (0.1 mg/mL PBS) 10 mL
Repeats: 20 times   

2. Purification of mouse IgG1 from ascitic fluid

Mouse ascitic fluid was 2-fold  diluted with PBS, applied to gel and shaked for 2 hrs. After washing, IgG was eluted at pH 2.8. Ab-Capcher ExTra shows 5.1-fold higher amount of IgG binding than that of Protein G gel and 30% higher than that of Ab-Capcher.

Gel volume: 50 μL/column
Sample: 1 mL of mouse ascitic fluid, x2 diluted with PBS
Incubation with gel: 2 hrs

 

3. Purification of rat IgGs from rat serum
Rat IgGs were purified from rat serum with approximately 95% recovery, showing approximately 20% higher than that of Ab-Capcher.

Frequently Asked Questions

Product List

Product Name Cat# Quantity Price

Ab-Capcher ExTra

PTN-P-003-2 2ML

¥ 30,800
$ 411
€ 308

Ab-Capcher ExTra

PTN-P-003-10 10ML

¥ 96,800
$ 1291
€ 968

Ab-Rapid SPiN Ex

PTN-P-014-10 10COLUMN

¥ 23,400
$ 312
€ 234

Ab-Rapid SPiN Ex

PTN-P-014-5-1 2COLUMN

¥ 59,800
$ 798
€ 598

Ab-Rapid PuRe Ex

PTN-P-015-2 2COLUMN

¥ 20,900
$ 279
€ 209

Ab-Rapid PuRe Ex

PTN-P-015-10 10COLUMN

¥ 83,600
$ 1115
€ 836

To be used for research only. DO NOT use for human gene therapy or clinical diagnosis.