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Table 2 Assay conditions such as cell lines, cell densities, corresponding ligands, incubation time with compounds, and materials for reporter gene assays in pathway profiling

From: Tubulin is a molecular target of the Wnt-activating chemical probe

Cellular signal

Cell lines

Cell density (cells/well)

Ligand

Incubation time with compounds

Original materials or references

cAMP response element (CRE) signal

HEK293T

5,000

Forskolin (1 μM)

5 h

pGL4.29 (Promega)

Nuclear factor of activated T-cells (NFAT) signal

HEK293T

5,000

Ionomycin (1 μM) PMA (10 ng/mL)

5 h

pGL4.30 (Promega)

Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) signal

HEK293T

10,000

TNFα (3 ng/mL)

20 h

pGL4.32 (Promega)

Serum response element (SRE) signal

HEK293T

20,000

FBS (15 %) PMA (30 ng/mL)

20 h

pGL4.33 (Promega)

Serum response factor (SRF) signal

HEK293T

20,000

FBS (15 %)

5 h

pGL4.34 (Promega)

p53 signal

HEK293T

10,000

Doxorubicin (1 μM)

20 h

[43]

E2F signal

HEK293T

20,000

FBS (15 %)

20 h

[44]

Activating transcription factor 6 signal

HEK293T

5,000

Thapsigargin (30 nM)

5 h

[45]

Hedgehog signal

NIH3T3

7,500

mouse sonic hedgehog

20 h

[46]

Hypoxia-inducible factor 1 (HIF1) signal

HEK293T

10,000

hypoxia

20 h

[47]

Nuclear factor erythroid 2-related factor 2 (Nrf2) signal

HEK293T

5,000

tert-butylhydroquinone (20 μM)

20 h

pGL4.37 (Promega)

SMAD signal

HEK293T

10,000

TGFβ (0.2 ng/mL)

20 h

[48]

Wnt signal

HEK293T

10,000

Wnt3a

20 h

[49]

Wnt signal

SW480

10,000

no ligand (constitutive active)

20 h

[49]

IL17 signal

Jurkat

5,000

Ionomycin (400 nM) PMA (4 ng/mL)

5 h

[50]