The recommended protocol for using Alizarin Red S involves a pH 4.2 method. Prepare the solution by adding 2 grams of Alizarin Red S to 100 ml of deionized water and mix well. Adjust the pH to 4.1 to 4.3 using a 0.5% ammonium hydroxide solution. It is important to use a relatively new bottle of ammonium hydroxide. If the pH is initially more alkaline than 4.2, use HCl to lower it below 4.2, then adjust with ammonium hydroxide. The pH and presence of ammonium ions are critical. Yellow background staining is acceptable, and if there is no calcium in the tissue, the staining will appear yellow. Ensure a positive control is used that has not been exposed to decalcifying agents, as these will remove calcium from tissue sections.
A5533
Alizarin Red S
certified by the Biological Stain Commission, powder
Synonym(s):
3,4-Dihydroxy-9,10-dioxo-2-anthracenesulfonic acid sodium salt, Alizarin Carmine, Alizarin Red, water soluble, Alizarin sodium monosulfonate, Alizarin sulfonate sodium, Alizarinsulfonic acid sodium salt, Sodium alizarinsulfonate
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All Photos(2)
About This Item
Empirical Formula (Hill Notation):
C14H7NaO7S
CAS Number:
Molecular Weight:
342.26
Colour Index Number:
58005
Beilstein:
4117090
EC Number:
MDL number:
UNSPSC Code:
12171500
PubChem Substance ID:
NACRES:
NA.47
Pricing and availability is not currently available.
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Product Name
Alizarin Red S, certified by the Biological Stain Commission
grade
certified by the Biological Stain Commission
Quality Level
form
powder
concentration
0.015 g/L in 0.1 M NaOH
technique(s)
titration: suitable
color
dark red
yellow-orange to dark orange
solubility
H2O: 1 mg/mL
ε (extinction coefficient)
592-598 nm at 0.015 g/L in 0.1 M NaOH
application(s)
diagnostic assay manufacturing
hematology
histology
storage temp.
room temp
SMILES string
[Na+].Oc1c(O)c(cc2C(=O)c3ccccc3C(=O)c12)S([O-])(=O)=O
InChI
1S/C14H8O7S.Na/c15-11-6-3-1-2-4-7(6)12(16)10-8(11)5-9(22(19,20)21)13(17)14(10)18;/h1-5,17-18H,(H,19,20,21);/q;+1/p-1
InChI key
HFVAFDPGUJEFBQ-UHFFFAOYSA-M
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Application
Biochem/physiol Actions
Suitability
Certified for use in Dahl′s method for calcium
Storage Class Code
11 - Combustible Solids
WGK
WGK 3
Flash Point(F)
Not applicable
Flash Point(C)
Not applicable
Personal Protective Equipment
dust mask type N95 (US), Eyeshields, Gloves
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Impaired marrow osteogenesis is associated with reduced endocortical bone formation but does not impair periosteal bone formation in long bones of SAMP6 mice.
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Journal of Bone and Mineral Research, 20, 419-419 (2005)
Alizarin Red S. as a general optical reporter for studying the binding of boronic acids with carbohydrates.
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Phenol red inhibits chondrogenic differentiation and affects osteogenic differentiation of human mesenchymal stem cells in vitro.
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RGD peptides immobilized on a mechanically deformable surface promote osteoblast differentiation.
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On the history and mechanism of alizarin and alizarin red S stains for calcium.
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The Journal of Histochemistry and Cytochemistry, 17, 110-124 (1969)
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My Alizarin red stain is staining my cells yellow instead of red. How should I adjust the solution so that it stains the cells red?
1 answer-
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What is the procedure for preparing a staining solution for Alizarin Red S when using product A5533?
1 answer-
The Alizarin Red S dye is available as a powdered dye, with catalog number A5533 recommended (BSC certified pH 7.0). Dissolve 2 grams of Alizarin Red S in 100 ml of distilled water and adjusting the pH of the solution to pH 4.2. It's important to adjust the pH of the dilute ammonium hydroxide solution. If the initial pH is higher than pH 4.3, a little dilute HCL should be used to lower the pH, then adjust the pH back to a range of between pH 4.1 to pH 4.3. The ammonium ions are necessary for proper staining to occur.
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Can A5533 be utilized for fluorescent imaging, specifically for confocal imaging purposes?
1 answer-
The fluorescent properties of Alizarin Red S are not tested. As for its use in confocal microscopy, there are no applications mentioned.
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Hi, do you have a protocol for preparing the alizarin red s powder and for staining human osteoblast cells to detect mineralisation? Thank you
1 answer-
This product is not routinely tested for staining osteoblast cells. A general calcium staining protocol for use with paraffin-embedded samples is available below:
Calcium Staining protocol for use with Alizarin Red S
Fixation: 10 % Neutral Buffered Formalin or alcoholic formalin. Do not use calcium-containing fixatives
Embedding: Embed in paraffin and cut sections at 5 microns
Preparation of solutions:
Alizarin Red S, certified 2 grams
Distilled water 100 mL
Mix well and adjust the pH to 4.1 to 4.3 with a dilute ammonium hydroxide solution.
Procedure:
1. Deparaffinize, hydrate slides, and place slides in 50 % alcohol.
2. Rinse rapidly with distilled water.
3. Lay slides flat and cover with the alizarin red s solution.
4. Observe the reaction under a microscope and remove when a red-orange lake forms (usually 30 seconds to 5 minutes. The lake should be heavy, but not too diffused.
5. Shake off excess dye and blot, carefully.
6. Dehydrate in acetone for 10 to 20 seconds and in acetone-xylene (50:50) for 10 to 20 seconds.
7. Clear in xylene and mount in a synthetic mounting medium
Results:
Calcium deposits (except oxalate) will stain orange-red. Precipitate is birefringent.Alternatively, product TMS-008, Alizarin Red Staining Solution, has been tested for use with human and rodent osteocytes.
See the links below to review the product information, as well as an application note:
TMS-008 Alizarin-Red Staining Solution:
https://www.sigmaaldrich.com/product/mm/tms008Osteocyte Differentiation Application Note:
https://www.sigmaaldrich.com/technical-documents/technical-article/cell-culture-and-cell-culture-analysis/stem-cell-culture/mesenchymal-stem-cell-osteogenesisHelpful?
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This product can be used in a in vivo bone immunolabeling ? I see a work available for you in this site using the Alizarin Red (A5533) to study bone regeneration. They inject the alizarin red S intraperitoneal some days before to sacrifice the animals.
1 answer-
This product has not been tested for in vivo or in vitro applications. Suitability would have to be determined by the end-user. Please see the link below to a publication that may be helpful:
https://pubmed.ncbi.nlm.nih.gov/15746986/Helpful?
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Can you please help clarify the difference between Alizarin-Red (TMS-008) and Alizarin Red S (A5533)? Also, in what situations would one be more suitable than the other?
1 answer-
Product A5533 is an Alizarin Red S powder while the TMS-008 is simply the 40 mM solution in water. Both products are used in the staining of cartilage and bone tissue. The A5533 is a Sigma-Aldrich brand and the TMS-008 is an EMD Millipore brand. Functionally, these products may be used for the same application.
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Is there an available excitation / emission specrtrum for this Alizarin Red S for fluorescence studies?
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Various publications differ somewhat. Most sources report an excitation range of 530-560nm and the emission is expected to be 580nm. Significant shifts from these values can be found in the literature from due to varying conditions - pH, solvent, and sample type.
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I see that Product A5533, Alizarin Red S, was tested according to the Dahl method at pH 7.0. Can the dye also be used in the McGee-Russell procedure at pH 4.2?
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Yes. The dye can be used at a variety of pH values.
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What is the Department of Transportation shipping information for this product?
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Transportation information can be found in Section 14 of the product's (M)SDS.To access the shipping information for this material, use the link on the product detail page for the product.
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What are the pH transition ranges for Product A5533, Alizarin Red S?
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Although the pH transition ranges are not determined on a lot to lot basis, the solution changes from yellow to red in the range of 3.5-6.5, and from the orange to violet at pH 9.4-12. On a lot to lot basis there can be considerable difference in regards to color and pH ranges.
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