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Human c-Myc Cell-based Kit HTRF®

The Human c-Myc Cell-based kit is designed for the rapid detection of human c-Myc proto-oncogene in cells.
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  • Ready-to-use Ready-to-use
  • Highly specific Highly specific
  • Faster and more convenient than ELISA Faster and more convenient than ELISA
The Human c-Myc Cell-based kit is designed for the rapid detection of human c-Myc proto-oncogene in cells.
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Overview

Human c-Myc is a multifunctional transcription factor protein that plays a role in cell cycle progression, apoptosis, and cellular transformation. c-Myc also functions to regulate global chromatin structure by regulating histone acetylation. The kit is intended for the specific and fast quantification of c-Myc proto-oncogene in cells.

Benefits

  • FASTER ALTERNATIVE TO ELISA OR WB
  • HIGHLY SPECIFIC
  • READY-TO-USE KIT

Assay principle

The Human c-Myc is a sandwich immunoassay involving two specific anti-human c-Myc antibodies, respectively labelled with Europium Cryptate (donor) and d2 (acceptor). The intensity of the signal is proportional to the concentration of c-Myc present in the sample. The protocol is optimized for a 384-well plate format, but can easily be further miniaturized or upscaled. Only low sample volumes are needed. The detection reagents may be pre-mixed and added in a single dispensing step for direct detection. No washing is needed at any step.
Human c-Myc assay principle

Assay Protocol

Two-plate assay protocol The human c-Myc assay protocol is described here. Cells are plated, stimulated, and lysed in the same 96-well culture plate. Lysates are then transferred to the assay plate for the detection of human c-Myc by HTRF reagents. This protocol enables the cells' viability and confluence to be monitored. The antibodies labelled with HTRF fluorophores may be pre-mixed and added in a single dispensing step to further streamline the assay procedure. The assay detection can be run in 96- to 384-well plates by simply resizing each addition volume proportionally.
Human c-Myc assay protocol

HTRF assay compared to Western Blot

HeLa cells were grown in a T175 flask at 37°C - 5% CO2 for 48 h. After medium removal, the cells were lysed with 3 mL of supplemented lysis buffer for 45 min at RT. Serial dilutions of the cell lysate were performed in the supplemented lysis buffer and 10 µL of each dilution were dispensed and analyzed side-by-side by Western-blot and by HTRF. By using HTRF c-Myc only 1,000 cells are sufficient for minimal signal detection while 4,000 cells are needed for a Western Blot signal. The HTRF c-Myc assay is at least 4-fold more sensitive than the Western Blot.
c-Myc protein level comparison HTRF vs Western Blot

c-Myc detection in several human cell lines

100,000 human MCF7, A549, HeLa, HTC116 and HEK293T cells were plated in 96-well plates in cell culture medium and incubated for 24h at 37°C - 5% CO2. Cell culture medium was then removed and cells were lysed with 50 µL of lysis buffer for 30 min at RT under gentle shaking. 10 µL of lysate was transferred into a 384-well sv white microplate, and 10 µL of the HTRF c-Myc detection reagents were added. The HTRF signal was recorded after an overnight incubation time.
c-Myc detection in various human cell lines

Bromodomain interaction inhibitor effect

40,000 HeLa cells were plated in 96 well plate in cell culture medium and incubated for 24h at 37°C - 5% CO2. After incubating with increasing concentrations of JQ1 for 24 hours, the medium was removed and cells were lysed with 50 µL of lysis buffer for 45 min at RT under gentle shaking. 10 µL of lysate were transferred into a 384-well sv white microplate and 10 µL of the HTRF c-Myc detection reagents were added. The HTRF signal was recorded after an overnight incubation. 

Bromodomain interaction inhibitor effect on c-Myc detection

Histone deacetylase (HDAC) inhibitor effect

40,000 HeLa cells were plated in 96-well plates in cell culture medium, and incubated for 24h at 37°C - 5% CO2. After incubating with increasing concentrations of TSA for 24 hours, the medium was removed and cells were lysed with 50 µL of lysis buffer for 45 min at RT under gentle shaking. 10 µL of lysate were transferred into a 384-well sv white microplate and 10 µL of the HTRF c-Myc detection reagents were added. The HTRF signal was recorded after an overnight incubation. 

Histone deacetylase inhibitor effect on c-Myc detection

Cell growth inhibitors effect

10,000 HeLa or A549 cells were plated in 96-well plates in cell culture medium and incubated for 24h at 37°C - 5% CO2. After incubating with increasing concentrations of inhibitors (U0126, Methotrexate and Gemcitabine) for 72 hours, the medium was removed and cells were lysed with 50 µL of lysis buffer for 45 min at RT under gentle shaking. 10 µL of lysate were transferred into a 384-well sv white microplate and 10 µL of the HTRF c-Myc detection reagents were added. The HTRF signal was recorded after an overnight incubation.
Cell growth inhibitor effect on c-Myc detection

Product Insert c-Myc (h) Kit / 63ADK053PEG-63ADK053PEH

63ADK053PEG-63ADK053PEH - Product Insert

HTRF Product Catalog

All your HTRF assays in one document! - Catalog

A guide to Homogeneous Time Resolved Fluorescence

General principles of HTRF - Guides

How HTRF compares to Western Blot and ELISA

Get the brochure about technology comparison. - Brochures

Inhibition of FGFR Signaling Pathways Studied in Cancer Cell Lines Using HTRF

The knowledge and data necessary to understand the inhibition value of FGFR receptors involved in human cancers. HTRF technology enables the monitoring of the activity of the AZD4547 inhibitor on cell signal transduction in 3 cancer cell lines. - Application Notes

Safety Data Sheet c-Myc (h) Kit / 63ADK053PEH

63ADK053PEH - Safety Data Sheet

Plate Reader Requirement

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