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  5. Thermo Scientific™ Pierce™ Nickel Coated Plates

Thermo Scientific™ Pierce™ Nickel Coated Plates

Catalog No. PI15442
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Color:
Black
Clear
White
Format:
Standard
Strip Plate
Detection Method:
Chemiluminescence
Colorimetric
Fluorescence
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Catalog No. Color Format Detection Method
PI15442 Clear Standard Colorimetric
PI15342 Black Standard Fluorescence
PI15142 Clear Strip Plate Colorimetric
PI15242 White Standard Chemiluminescence
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Catalog No. PI15442 Supplier Thermo Scientific™ Supplier No. 15442
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Description

Thermo Scientific™ Pierce Nickel Coated Plates provide a simple format to bind His-tagged fusion proteins for ELISA and other plate-based assays involving protein interactions of expressed recombinant proteins.

These Ni(2+) chelate-coated plates are ideal for analyzing polyhistidine-tagged fusion proteins by ELISA-based methods. Proteins that contain a succession of several histidine residues at the amino or carboxyl terminus have a strong binding affinity for metal. Bacterial lysates containing polyhistidine-tagged fusion proteins can be added directly to the plates without the need for blocking. The clear, white or black plates can be used with colorimetric, chemiluminescent or fluorescent detection methods, respectively.

Thermo Scientific Pierce Nickel Coated White 96-Well Plates provide a simple format to bind His-tagged fusion proteins for ELISA and other plate-based assays involving protein interactions of expressed recombinant proteins. These white plates can be used with chemiluminescent detection methods.

Features of Pierce Nickel Coated Plates include:
• Ni2+ activated surface enables metal-chelate binding of His-tagged proteins
• Detergents used to lyse cells don't inhibit binding to activated plates as they do with plain polystyrene
• Better binding for sensitive assays compared to other commercially available nickel-activated plates
• Detection limit: 1 ng of polyhistidine fusion protein
• Binding capacity: approx. 9 pmol His-tagged protein (27 kDa) per well
• Activation level: 200 μL

These Ni(2+) chelate-coated plates are ideal for analyzing polyhistidine-tagged fusion proteins by ELISA-based methods. Proteins that contain a succession of several histidine residues at the amino or carboxyl terminus have a strong binding affinity for metal. Bacterial lysates containing polyhistidine-tagged fusion proteins can be added directly to the plates without the need for blocking.

Specifications

Binding Property Standard Capacity, Affinity
Format Standard
Content And Storage Upon receipt store plates at 4°C in unopened pouches. Once opened, place unused plates in a resealable bag with desiccant and store at 4°C.
Detection Method Colorimetric
Packaging 5 Plates
Product Line Pierce
Color Clear
Product Type Microplate
Material Polystyrene
Volume (Metric) Well 200 μL
No. of Wells 96
Surface Treatment Divalent Nickel, Metal Chelate
Plate Blocking Blocker™ BSA Buffer
Quantity 5 plates
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Frequently Asked Questions (FAQs)

What is the binding capacity for Pierce Nickel Coated Plates?

The binding capacity for Pierce Nickel Coated Plate is approximately 9 pmol His-tagged protein (27 kDa) per well.
Why should I use ultra-pure water for washing Pierce Nickel Coated Plates?

The water used for washing of microplates as well as all assay reagents, must be of absolutely ultra-pure quality. This means that no metal ions should be present in the water, as this will bind to the His-tag and thereby decrease the binding of the fusion protein to the nickel-chelate complex.
Is it possible to elute the bound His-tagged fusion protein/peptide from the surface of Pierce Nickel Coated Plates?

Yes. For instance, use imidazole in concentrations >500 mM in Tris at pH 7.5 or a high concentration of EDTA.
With Pierce Nickel Coated Plates, can any reagent interfere with the binding between the nickel-chelate complex and the His-tagged fusion biomolecule?

Ionic detergents (e.g., SDS) will interfere with the binding as well as high concentrations of chelating reagents like EDTA, EGTA, and very strong electron donors like metal ions.
Can I use Pierce Nickel Coated Plates to measure the amount of bound His-tagged fusion protein/peptide?

To determine the approximate amount of bound His-tagged fusion protein/peptide, a standard curve of a previously purified preparation can be applied.
With Pierce Nickel Coated Plates, what is the detection limit for proteins?

The detection limit for each protein depends on the assay system used (e.g., primary and secondary antibody, incubation time, detection reagent), the accessibility of the His-tagged fusion biomolecule, and the size of the protein (large proteins bind with a low density). Using a 25 kDa 6xHis-tagged protein, we have observed a detection limit of 1.0 ng per well (100 µL).
What is the stability of the Pierce Nickel Coated Plates?

The Pierce Nickel Coated Plates are extremely stable over a long period of time, if stored at room temperature (20-25 degrees C). We guarantee 12 mths of shelf life in unopened foil package after the date of shipment.
Should I optimize the concentration of the His-tagged protein/peptide for use with Pierce Nickel Coated Plates?

Yes. When setting-up the assay, the concentration of the His-tagged protein/peptide should be determined by preparing a titration. We recommend using a solution of the His-tagged fusion protein/peptide diluted in KCl, in a concentration ranging from 0.01-10 µg/mL. Hereafter, the primary and secondary antibodies should be optimized according to the manufacturer's recommendations.
Why do you recommend pre-washing Pierce Nickel Coated Plates?

We recommend pre-washing Pierce Nickel Coated Plates with PBS containing 0.05 % (v/v) Tween-20 surfactant to saturate the plate surface. This minimizes the non-specific binding during the assay and provides a low background.
In Pierce Nickel Coated Plates, is the interaction between the nickel-chelate complex and histidine pH-dependent?

Yes. The interaction is pH-dependent, and binding is best obtained at neutral pH or slightly above neutral pH. We recommend using 10 mM KCl, which has a neutral pH, as a coupling buffer.
Can the Pierce Nickel Coated Plates bind proteins containing less than six neighboring His amino acids?

Due to steric hindrance, optimal performance is normally observed if the fusion protein contains more than four neighboring histidine amino acids located at the terminal end. In some cases the histidine amino acids do not need to be placed next to each other - just be sure that the 3D structure fold them next to each other (e.g., the HAT fusion protein).
Which type of molecules can I use with Pierce Nickel Coated Plates?

The Pierce Nickel Coated Plates can via a simple one-step protocol bind all types of His-tagged fusion proteins/peptides. This includes purified as well as crude cell lysates containing His-tagged fusion proteins/peptides.