Hoechst 33342 (Synonyms: bisBenzimide H 33342; HOE 33342)

Name: Hoechst 33342
Brand: MedChemExpress
SKU: HY-15559
Rating: 5.0 (68 reviews)

Cat. No.: HY-15559 Purity: 99.94%: FAQs
Data Sheet SDS COA Handling Instructions

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Hoechst 33342 is a marker dye in Hoechst series. Hoechst is A live nuclear marker dye. Hoechst binds to the grooves in the DNA double strand, which tends to be A/ T-rich DNA strand. Although it binds to all nucleic acids, the A/ T-rich double strand DNA significantly enhances fluorescence intensity Therefore,Hoechst dye can be used for living cell labeling. The fluorescence intensity of Hoechst dye increases with the increase of pH of solution.

For research use only. We do not sell to patients.

Hoechst 33342 Chemical Structure

CAS No. : 23491-52-3

Size	Price	Stock	Quantity
50 mg	USD 50	In-stock	Estimated Time of Arrival: December 31
100 mg	USD 85	In-stock	Estimated Time of Arrival: December 31
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Based on 68 publication(s) in Google Scholar

Other Forms of Hoechst 33342:

58 Publications Citing Use of MCE Hoechst 33342

: Hoechst 33342 purchased from MedChemExpress. Usage Cited in: Bioact Mater. 2022 Mar 17;18:91-103. [Abstract]; The fluorescent images of HUVECs stained with NO fluorescent probe (DAF-FM DA) and Hoechst 33342 after being treated with UCATS under dark conditions or 808 nm NIR irradiation.

: Hoechst 33342 purchased from MedChemExpress. Usage Cited in: Chem Eng J. 15 December 2022, 138004.; Apoptotic cells in four groups were stained with Hoechst 33342 (for 10 min) reflected by bright blue nuclei, including hP-MSCs cultured on noncoated, L-Gel-coated (10 nM), and D-Gel coated (10 nM) plates under exposure of 500 μM H2O2 for 5 h except for the normal group, and PBS as control.

: Hoechst 33342 purchased from MedChemExpress. Usage Cited in: J Lipid Res. 2022 May;63(5):100207. [Abstract]; complete medium containing 1 μg/mL BODIPY 500/510 for 3 h and then 2 μg/mL Hoechst 33342 for an additional 2 min.

: Hoechst 33342 purchased from MedChemExpress. Usage Cited in: Theranostics. 2021 Mar 4;11(10):4655-4671. [Abstract]; Hoechst 33342 (10 µg/mL) is added to the medium of cultured megakaryocytes for 2 h at 37°C. Cells are stained with the directly coupled monoclonal antibodies PE-anti-CD41 and APC-anti-Mpl for 30 min at 4°C. Ploidy is measured in the CD41+Mpl+ cell population.

: Hoechst 33342 purchased from MedChemExpress. Usage Cited in: J Cell Biol. 2021 Oct 4;220(10):e202011153. [Abstract]; Chromatin is briefly counterstained with Hoechst 33342 (5 µg/mL) before imaging.

: Hoechst 33342 purchased from MedChemExpress. Usage Cited in: Aging Cell. 2021 Jul;20(7):e13382. [Abstract]; BM cells are incubated with 5 μg/mL Hoechst 33342 in Hank balanced salt solution containing 20 mM HEPES, 5 mM glucose, and 10% fetal bovine serum at 37°C for 45 min, followed by an additional 45 min of incubation with 1 μg/ml Pyronin Y.

: Hoechst 33342 purchased from MedChemExpress. Usage Cited in: Stem Cell Res Ther. 2021 Jan 22;12(1):77. [Abstract]; To evaluate the anti-apoptosis property of extracellular vesicles from hP-MSCs, the Hoechst 33342 (for 10 min) is used to distinguish the apoptotic cells in the three different groups.

: Hoechst 33342 purchased from MedChemExpress. Usage Cited in: Acta Biomater. 2020 Sep 1;113:289-304. [Abstract]; Hoechst 33342 staining (for 6 h) is used to distinguish apoptotic changes in H2O2-treated hP-MSCs.

: Hoechst 33342 purchased from MedChemExpress. Usage Cited in: ACS Appl Mater Interfaces. 2018 Sep 12;10(36):30081-30091. [Abstract]; Hoechst 33342 staining shows the survival of HUVECs administrated 100 μg/mL exosomes or CS-Exo upon exposure to 400 μM H₂O₂.

: Hoechst 33342 purchased from MedChemExpress. Usage Cited in: Chinese Pharmacological Bulletin. 2018 May; 34(5): 620-626.; PC12 cells are treated with 1.5 mM Methylglyoxal (MG) for 24 h before pretreatment with Butein (2.5, 5 μM) for 1 h. Propidium iodide (PI) and Hoechst 33342 are used to determine cell apoptosis.

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Biological Activity
Protocol
Purity & Documentation
References
Customer Review

Description

IC₅₀ & Target

Dye reagent^[1]
DNA Stain^[1]

Cellular Effect

Cell Line	Type	Value	Description	References
A549	IC₅₀	0.98 μg/mL Compound: 9	Cytotoxicity against human A549 cells assessed as reduction in cell survival after 24 hrs by resazurin assay Cytotoxicity against human A549 cells assessed as reduction in cell survival after 24 hrs by resazurin assay	[PMID: 27301676]
BEAS-2B	IC₅₀	1.95 μg/mL Compound: 9	Cytotoxicity against human BEAS2B cells assessed as reduction in cell survival after 24 hrs by resazurin assay Cytotoxicity against human BEAS2B cells assessed as reduction in cell survival after 24 hrs by resazurin assay	[PMID: 27301676]
DU-145	IC₅₀	4.25 μM Compound: Hoechst 33342	Cytotoxicity against human DU145 cells after 48 hrs by sulforhodamine B assay Cytotoxicity against human DU145 cells after 48 hrs by sulforhodamine B assay	[PMID: 25083189]
HEK293	IC₅₀	18 μM Compound: Hoechst 33342	Inhibition of human liver OATP1B3 expressed in HEK293 Flp-In cells assessed as reduction in [3H]E17-betaG uptake incubated for 5 mins by scintillation counting Inhibition of human liver OATP1B3 expressed in HEK293 Flp-In cells assessed as reduction in [3H]E17-betaG uptake incubated for 5 mins by scintillation counting	[PMID: 22541068]
HEK293	IC₅₀	19 μM Compound: Hoechst 33342	Inhibition of human liver OATP1B1 expressed in HEK293 Flp-In cells assessed as reduction in E17-betaG uptake by scintillation counting Inhibition of human liver OATP1B1 expressed in HEK293 Flp-In cells assessed as reduction in E17-betaG uptake by scintillation counting	[PMID: 22541068]
HEK293	IC₅₀	51 μM Compound: Hoechst 33342	Inhibition of human liver OATP2B1 expressed in HEK293 Flp-In cells assessed as reduction in [3H]E3S uptake incubated for 5 mins by scintillation counting Inhibition of human liver OATP2B1 expressed in HEK293 Flp-In cells assessed as reduction in [3H]E3S uptake incubated for 5 mins by scintillation counting	[PMID: 22541068]
HeLa	IC₅₀	0.3 μM Compound: Hoechst 33342	Cytotoxicity against human HeLa cells after 72 hrs by MTT assay Cytotoxicity against human HeLa cells after 72 hrs by MTT assay	[PMID: 21186067]
MCF7	IC₅₀	0.5 μM Compound: Hoechst 33342	Cytotoxicity against human MCF7 cells after 72 hrs by MTT assay Cytotoxicity against human MCF7 cells after 72 hrs by MTT assay	[PMID: 21186067]
U-87MG ATCC	IC₅₀	4.2 μM Compound: Hoechst 33342	Cytotoxicity against human U87 cells after 72 hrs by MTT assay Cytotoxicity against human U87 cells after 72 hrs by MTT assay	[PMID: 21186067]

In Vitro

General Protocol
Preparation of Hoechst working solution
1.1 Preparation of the stock solution
Dissolve 10 mg of in 5 mL DMSO
Note: It is recommended to store the stock solution at 4℃ or -20℃ away from light and avoid repetitive freeze-thaw cycles.
1.2 Preparation of Hoechst working solution
Dilute the stock solution in serum-free cell culture medium or PBS to obtain final concentration 10 μg/mL Hoechst working solution.
Note: Please adjust the concentration of Hoechst working solution according to the actual situation.
1.Cell staining
2.1 Suspension cells（6-well plate）
a. Centrifuge at 1000 g at 4℃ for 3-5 minutes and then discard the supernatant. Wash twice with PBS, 5 minutes each time. The cell density is 1×10⁶/mL.
b. Add 1 mL of working solution, and then incubate at room temperature for 3-10 minutes.
c. Centrifuge at 400 g at 4℃ for 3-4 minutes and then discard the supernatant.
d. Wash twice with PBS, 5 minutes each time.
e. Resuspend cells with serum-free cell culture medium or PBS. Observation by fluorescence microscopy or flow cytometry.
2.2 Adherent cells
a. Culture adherent cells on sterile coverslips.
b. Remove the coverslip from the medium and aspirate excess medium.
c. Add 100 μL of working solution, gently shake it to completely cover the cells,and then incubate at room temperature for 3-10 minutes.
d. Wash twice with medium, 5 minutes each time.Observation by fluorescence microscopy or flow cytometry.

Precautions
1. Please adjust the concentration of Hoechst working solution according to the actual situation.
2. This product is for R&D use only, not for drug, household, or other uses.
3. For your safety and health, please wear a lab coat and disposable gloves to operate.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

Molecular Weight

452.55

Formula

C₂₇H₂₈N₆O

CAS No.

23491-52-3

Appearance

Solid

Color

Brown to breen

Emission (Em)

452

Excitation (Ex)

356

SMILES

CN1CCN(C2=CC=C3C(N=C(C4=CC=C5C(N=C(C6=CC=C(OCC)C=C6)N5)=C4)N3)=C2)CC1

Shipping

Room temperature in continental US; may vary elsewhere.

Storage

4°C, protect from light

*In solvent : -80°C, 6 months; -20°C, 1 month (protect from light)

Solvent & Solubility

In Vitro:

DMSO : 6.25 mg/mL (13.81 mM; ultrasonic and warming and heat to 60°C; Hygroscopic DMSO has a significant impact on the solubility of product, please use newly opened DMSO)

H₂O : < 0.1 mg/mL (insoluble)

Preparing
Stock Solutions

Concentration Solvent Mass	1 mg	5 mg	10 mg

1 mM	2.2097 mL	11.0485 mL	22.0970 mL
5 mM	0.4419 mL	2.2097 mL	4.4194 mL
10 mM	0.2210 mL	1.1049 mL	2.2097 mL

View the Complete Stock Solution Preparation Table

* Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (protect from light). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

Molarity Calculator
Dilution Calculator

Mass (g) = Concentration (mol/L) × Volume (L) × Molecular Weight (g/mol)

Concentration _(start) × Volume _(start) = Concentration _(final) × Volume _(final)

This equation is commonly abbreviated as: C₁V₁ = C₂V₂

Concentration _(start)

Volume _(start)

Concentration _(final)

Volume _(final)

In Vivo:

Select the appropriate dissolution method based on your experimental animal and administration route.

For the following dissolution methods, please ensure to first prepare a clear stock solution using an In Vitro approach and then sequentially add co-solvents:
To ensure reliable experimental results, the clarified stock solution can be appropriately stored based on storage conditions. As for the working solution for in vivo experiments, it is recommended to prepare freshly and use it on the same day.
The percentages shown for the solvents indicate their volumetric ratio in the final prepared solution. If precipitation or phase separation occurs during preparation, heat and/or sonication can be used to aid dissolution.

Protocol 1

Add each solvent one by one: 10% DMSO 40% PEG300 5% Tween-80 45% Saline
Solubility: ≥ 0.5 mg/mL (1.10 mM); Clear solution

This protocol yields a clear solution of ≥ 0.5 mg/mL (saturation unknown).
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (5.0 mg/mL) to 400 μL PEG300, and mix evenly; then add 50 μL Tween-80 and mix evenly; then add 450 μL Saline to adjust the volume to 1 mL.
Preparation of Saline: Dissolve 0.9 g sodium chloride in ddH₂O and dilute to 100 mL to obtain a clear Saline solution.
Protocol 2

Add each solvent one by one: 10% DMSO 90% (20% SBE-β-CD in Saline)
Solubility: ≥ 0.5 mg/mL (1.10 mM); Clear solution

This protocol yields a clear solution of ≥ 0.5 mg/mL (saturation unknown).
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (5.0 mg/mL) to 900 μL 20% SBE-β-CD in Saline, and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C, storage for one week): 2 g SBE-β-CD powder is dissolved in 10 mL Saline, completely dissolve until clear.
Protocol 3

Add each solvent one by one: 10% DMSO 90% Corn Oil
Solubility: ≥ 0.5 mg/mL (1.10 mM); Clear solution

This protocol yields a clear solution of ≥ 0.5 mg/mL (saturation unknown). If the continuous dosing period exceeds half a month, please choose this protocol carefully.
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (5.0 mg/mL) to 900 μL Corn oil, and mix evenly.

In Vivo Dissolution Calculator

Please enter the basic information of animal experiments:

Dosage

mg/kg

Animal weight
(per animal)

Dosing volume
(per animal)

μL

Number of animals

Recommended: Prepare an additional quantity of animals to account for potential losses during experiments.

Please enter your animal formula composition:

DMSO +

Tween-80 +

Saline

Recommended: Keep the proportion of DMSO in working solution below 2% if your animal is weak.

The co-solvents required include: DMSO, . All of co-solvents are available by MedChemExpress (MCE). , Tween 80. All of co-solvents are available by MedChemExpress (MCE).

Calculation results:

Working solution concentration: mg/mL

Method for preparing stock solution: mg drug dissolved in μL DMSO (Stock solution concentration: mg/mL).

*In solvent : -80°C, 6 months; -20°C, 1 month (protect from light)

The concentration of the stock solution you require exceeds the measured solubility. The following solution is for reference only. If necessary, please contact MedChemExpress (MCE).

Method for preparing in vivo working solution for animal experiments: Take μL DMSO stock solution, add μL . μL , mix evenly, next add μL Tween 80, mix evenly, then add μL Saline.

If the continuous dosing period exceeds half a month, please choose this protocol carefully.

Please ensure that the stock solution in the first step is dissolved to a clear state, and add co-solvents in sequence. You can use ultrasonic heating (ultrasonic cleaner, recommended frequency 20-40 kHz), vortexing, etc. to assist dissolution.

Purity & Documentation

Purity: 99.94%

Select Batch:

Data Sheet (276 KB) SDS (393 KB)

COA (245 KB) HNMR (414 KB) RP-HPLC (252 KB) LCMS (268 KB)

Handling Instructions (2659 KB)

Dyeing Example

References

[1]. Chazotte B. Labeling nuclear DNA with hoechst 33342. Cold Spring Harb Protoc. 2011 Jan 1;2011(1):pdb.prot5557. [Content Brief]

Cell Assay
^[1]

Labeling Nuclear DNA with Hoechst 33342^[1] Step 1, Dilute the Hoechst stock solution 1:100 in H₂O for use in labeling. Step 2, Aspirate the cell medium from cells grown on coverslips. Rinse the cells three times with PBS⁺. Step 3, Incubate the cells in the Hoechst labeling solution (from Step 1) for 10-30 min at room temperature. Step 4, Aspirate the labeling solution. Rinse the cells three times in PBS⁺. Step 5, Mount the coverslips. Step 6, Image the cells (λex ~353 nm, λem ~483 nm for Hoechst 33342)^[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

References

[1]. Chazotte B. Labeling nuclear DNA with hoechst 33342. Cold Spring Harb Protoc. 2011 Jan 1;2011(1):pdb.prot5557. [Content Brief]

Complete Stock Solution Preparation Table

Optional Solvent	Concentration Solvent Mass	1 mg	5 mg	10 mg	25 mg

DMSO	1 mM	2.2097 mL	11.0485 mL	22.0970 mL	55.2425 mL
	5 mM	0.4419 mL	2.2097 mL	4.4194 mL	11.0485 mL
	10 mM	0.2210 mL	1.1049 mL	2.2097 mL	5.5243 mL

Hoechst 33342 Related Classifications

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Do most proteins show cross-species activity?
Species cross-reactivity must be investigated individually for each product. Many human cytokines will produce a nice response in mouse cell lines, and many mouse proteins will show activity on human cells. Other proteins may have a lower specific activity when used in the opposite species.

Keywords:

Hoechst 3334223491-52-3bisBenzimide H 33342 HOE 33342Hoechst33342Hoechst-33342bisBenzimide H33342bisBenzimide H-33342HOE33342HOE 33342HOE-33342AutophagyHoechstDNAcellular dyeblue fluorescenceInhibitorinhibitorinhibit

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Hoechst 33342 (Synonyms: bisBenzimide H 33342; HOE 33342)

Customer Review

Other Forms of Hoechst 33342:

Hoechst 33342 Related Antibodies

58 Publications Citing Use of MCE Hoechst 33342

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•Related Small Molecules:

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Customer Review

Hoechst 33342 Related Antibodies

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