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Biocytin C2 maleimide

Chemical structure for Biocytin C2 maleimide
Chemical structure for Biocytin C2 maleimide
Chemical structure for Biocytin C2 maleimide
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Physical properties
Molecular weight608.63
SolventDMSO
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
StorageFreeze (< -15 °C)
UNSPSC12352200

OverviewpdfSDSpdfProtocol


CAS
1248344-33-3
Molecular weight
608.63
Biocytin maleimide readily reacts with thiol moieties of biopolymers to form thioether conjugate that is quite stable. This biocytin maleimide requires mild conjugation conditions. For example, pH of 5.5-8.5 is usually optimal for modifying cysteine residues, and air exposure of reaction solution should be minimized whenever possible to avoid the air oxidation of thiol substrates. Most conjugations are done at room temperature. However, either elevated or reduced temperature may be required for a particular labeling reaction. Reactions with this biotinylation reagent should be performed in buffers free of extraneous thiols (such as 6-mercaptoethanol, dithiothreitol and mercaptoethylamine). Proteins or peptides to be biotinylated by thiol-reactive reagents must have a free thiol group (SH) available.

Example protocol


SAMPLE EXPERIMENTAL PROTOCOL

Labeling Proteins with Biocytin C2 Malemide
  1. Dissolve your thiol-containing protein at concentration of 1 - 10 mg/mL (3 - 10 mg is the optimal labeling concentration) using PBS buffer (20 mM, pH 7.2).
  2. Dissolve the Biocytin C2 Maleimide in DMSO at a concentration of 5 - 10 mg/mL.
  3. Mix the Biocytin C2 Maleimide and protein solution at 2:1 molar ratio of biocytin/protein, and shake the reaction mixture at room temperature for 2 - 4 hours.
  4. Filter the reaction mixture through a protein spin column for 100 µg to 1 mg protein labeling reaction. If the reaction scale is larger than 1 mg, purify the conjugate using gel filtration on a properly sized Sephadex G-25 column.
  5. Collect the desired fractions for your immediate use or freeze dry them for future use.
Labeling Small Molecules with Biocytin C2 Malemide
  1. Dissolve Biocytin C2 Maleimide (10 - 15 mg/mL) and your thiol-containing molecule in DMSO at 1:1.2 molar ratio of biocytin/thiol-containing molecule.
  2. Stir the reaction mixture at room temperature for 2 - 4 hours.
  3. Purify the conjugate using HPLC (ammonium acetate/water and acetonitrile, pH 7.0).
  4. Collect and pool the desired fractions.
  5. Combine and freeze-dry the pooled fractions.

Calculators


Common stock solution preparation

Table 1. Volume of DMSO needed to reconstitute specific mass of Biocytin C2 maleimide to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.

0.1 mg0.5 mg1 mg5 mg10 mg
1 mM164.303 µL821.517 µL1.643 mL8.215 mL16.43 mL
5 mM32.861 µL164.303 µL328.607 µL1.643 mL3.286 mL
10 mM16.43 µL82.152 µL164.303 µL821.517 µL1.643 mL

Molarity calculator

Enter any two values (mass, volume, concentration) to calculate the third.

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Product Family


NameExcitation (nm)Emission (nm)Extinction coefficient (cm -1 M -1)Correction Factor (280 nm)
AMCA C2 Maleimide346434190000.153
EDANS C2 maleimide33645559000.107
DABCYL C2 maleimide---0.516
XFD647 C2 Maleimide *Same Structure to Alexa Fluor™ 647 C2 Maleimide*6506712390000.03

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References


View all 51 references: Citation Explorer
The anterograde and retrograde axonal transport of biotinylated dextran amine and biocytin in the nervous system of teleosts
Authors: Xue HG, Yang CY, Ito H.
Journal: Brain Res Brain Res Protoc (2004): 106
Correlative fluorescence and electron microscopy of biocytin-filled neurons with a preservation of the postsynaptic ultrastructure
Authors: Morozov Y, Khalilov I, Ben-Ari Y, Represa A.
Journal: J Neurosci Methods (2002): 81
Biocytin and biotin uptake into NB2a neuroblastoma and C6 astrocytoma cells
Authors: Baur B, Suormala T, Baumgartner ER.
Journal: Brain Res (2002): 111
A new quantitative analytical method of serum biotinidase activity using biocytin as a substrate and its clinical significance in Japan
Authors: Kumasaka K, Muratsugu M, Fukui T, Kimura M, Takagi Y, Hashizume N.
Journal: Clin Chim Acta (2001): 71
Biotin and biocytin uptake into cultured primary calf brain microvessel endothelial cells of the blood-brain barrier
Authors: Baur B, Baumgartner ER.
Journal: Brain Res (2000): 348
In vitro biocytin injection into perinatal mouse brain: a method for tract tracing in developing tissue
Authors: Chang SL, LoTurco JJ, Nisenbaum LK.
Journal: J Neurosci Methods (2000): 1
A method of combining biocytin tract-tracing with avidin-biotin-peroxidase complex immunocytochemistry for pre-embedding electron microscopic labeling in neonatal tissue
Authors: Erisir A, Aoki C.
Journal: J Neurosci Methods (1998): 189
A rapid method for combined laser scanning confocal microscopic and electron microscopic visualization of biocytin or neurobiotin-labeled neurons
Authors: Sun XJ, Tolbert LP, Hildebr and JG, Meinertzhagen IA.
Journal: J Histochem Cytochem (1998): 263
A radioiodinated biocytin derivative for in-vivo applications
Authors: Nakamoto Y, Sakahara H, Saga T, Yao Z, Zhang M, Sato N, Zhao S, Arano Y, Ono M, Saji H, Konishi J.
Journal: Nucl Med Commun (1998): 963
Biotinylated dextran amine and biocytin hydrochloride are useful tracers for the study of retinal projections in the frog
Authors: Scalia F, Galoyan SM, Eisner S, Harris E, Su W.
Journal: J Neurosci Methods (1997): 167