Blockers for interference in sandwich immunoassays
Animal IgG – Passive Blockers • Suited for mixed species assays (e.g. MAb/PAb) • Species of blocker must be the same as the host of the capture or detection antibody
TRU Block ™ – Active HAMA & RF Blocker • Suited for double mouse monoclonal assays • Removes HAMA, HA & RF interference
TRU Block ™ Ready 8001 TRU Block ™ Ultra 8000 8002 (no azide) TRU Block ™ A66800H TRU Block ™ 2 A66802H TRU Block ™ 3 A66803H
Interfering HA (e.g. HAMA) TRU Block
Mouse IgG A66186M (9-13 mg/ml) A66186M-NA (9-13 mg/ml, Azide Free) A66185M (50-55 mg/ml) A66189M (45-55 mg/ml, Proclin) N14010M Mouse Serum
Detection Antibody (mouse origin) Capture Antibody (mouse origin) Antigen
Goat IgG A66200H Rat IgG A64391R A41182R
Rabbit IgG A66100H Sheep IgG A66400S (70-77mg/mL, Liquid)
*Formulations differ in their ratio of various proprietary ingredients that confers unique blocking characteristics.
In double mouse monoclonal assays, a specific blocker is required to remove a particular type of HA interference called human anti-mouse antibodies (HAMA) and Rheumatoid Factor (RF). A HAMA blocker contains a specific binder directed against all types of heterophilic interference including HAMA and RF. Once bound to the interfering antibodies, TRU Block prevents further binding of HA to other assay components through steric hindrance. Active blockers can typically be used in lower concentrations than passive blocking reagents, which minimizes the reduction in assay signal commonly associated with passive blockers.
Passive blocking reagents work by preventing interfering antibodies from binding to the capture or detection antibodies by providing alternate binding sites. Animal IgG (e.g. Goat IgG) can only block one type of interference (e.g. human anti-goat antibodies) so typically more than one type must be used, depending on the host of both the capture and detection antibodies. Animal IgG must be added in excess concentration and the effectiveness depends on the affinity of interfering antibody for the animal IgG. RECOMMENDED CONCENTRATION: • 10x the concentration of the MAb/PAb being used in the assay (e.g. if 5μg/mL of Ab/conjugate, add 50μg/ mL Animal IgG). • Can be added to the sample or conjugate diluent but ideally should be in contact with the patient sample before incubation with the assay capture antibody.
RECOMMENDED CONCENTRATION: For best performance, TRU Block should be included as part of the sample or conjugate diluent, at a recommended concentration range:
Protein Concentration Application
Single-step dilution with recommended dilution of 1:1000 to 1:10
TRU Block ™ Ready
ELISA & LF
TRU Block ™ ULTRA Range: 24-26 mg/mL ELISA, CLIA & LF TRU Block ™ ULTRA (no azide) Range: 24-26 mg/mL ELISA, CLIA & LF TRU Block ™ Range: 24-26 mg/mL ELISA & CLIA TRU Block ™ 2 Range: 24-26 mg/mL CLIA & LF TRU Block ™ 3 24.3 mg/mL ELISA
Blockers Practical Guide
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