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Purpose
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Enzyme-Linked Immunosorbent Assay (ELISA) for the detection of enzymatic activities that specifically transforms prelamin A precursor into mature lamin A protein in cell samples, as an aid in research studies and characterization of several laminopathies
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Sample Type
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Cell Lysate
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Analytical Method
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Quantitative
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Detection Method
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Colorimetric
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Purification
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Prelamin A recombinant protein was expressed in E.coli and purified by ion exchange and nichel sepharose affinity chromatographies.
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Components
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- Microtiter strips (12 x 8 well strips) coated with prelamin A recombinant protein, blocked with BSA 2% and protected with gelatine. - Lysis buffer A: 60ml ready to use, with preservative. - Lysis buffer B: 8ml ready to use, with preservative. - 2X Activity buffer: 8ml without FPP, SAM, DTT - Farnesyl Pyrophosphate (FPP): (farnesyl group donor) 200μl - S-(5'-Adenosyl)-L-methionine chloride (SAM): (methyl group donor) 0,5mg - DTT 1M: 250μl - Wash Buffer 10X concentrate: 100ml 10X to dilute to 1000ml final volume with distilled water, with preservative - Antibodies diluent: 30ml ready to use, with preservative - Anti-prelamin A primary antibody (ANT0045): 30μg - Anti-lamin A primary antibody (ANT0072): 15μg - HRP-conjugated secondary antibody: 15μl - Chromogen Solution: 30ml Buffer C, 30μl Buffer C1, 2 ABTS tablets.
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Material not included
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Microplate reader capable of measuring absorbance at 415 nm Thermomixer, shaking water bath or rocking platform at 37 °C Precision pipettes and pipette tips Glass or plastic pipettes Deionized or distilled water Multi-channel pipette, semi-automated or automated microplate washer 1000 mL graduated cylinder for preparation of 1X Wash Buffer Vortex mixer Glass tubes
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