Fluorescein 5-isothiocyanate → 2-(6-Hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

HRID 1154688Notes

Reaction equation

HRID 1154688 reaction equation
[CH:1]1[C:6](N=C=S)=[CH:5][C:4]2[C:10]([O:12][C:13]3([C:23]4[CH:24]=[CH:25][C:26]([OH:28])=[CH:27][C:22]=4[O:21][C:15]4[CH:16]=[C:17]([OH:20])[CH:18]=[CH:19][C:14]3=4)[C:3]=2[CH:2]=1)=[O:11].[I-]>O.C(=O)([O-])[O-].[Na+].[Na+]>[CH:1]1[CH:6]=[CH:5][C:4]([C:10]([OH:12])=[O:11])=[C:3]([C:13]2[C:14]3[CH:19]=[CH:18][C:17]([OH:20])=[CH:16][C:15]=3[O:21][C:22]3[C:23]=2[CH:24]=[CH:25][C:26]([CH:27]=3)=[O:28])[CH:2]=1

Participants

5

Reactant 2

HCID 18730Fluorescein 5-isothiocyanateHCID 30165Iodide Ion

Product 1

HCID 33832-(6-Hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid

Reagents, catalysts & solvents 2

HCID 962WaterSolventHCID 10340Sodium CarbonateSolvent

Procedure

To 0.34 mg fluorescein isothiocyanate placed as a dry solid in a screw-capped test tube was added 64 micrograms of N,N,N-trimethylammoniumbenzene iodide in 25 microliters of water, 0.2 ml 1M sodium carbonate buffer, pH 9.5, and 17 mg of the antibody product obtained in Example 3 in 2 ml PBS. The reaction was continued for six hours on ice, after which the solution was dialyzed against 0.015M NaCl, 0.01M sodium phosphate. This solution was applied to a DEAE-cellulose column equilibrated in the same buffer. Step-wise increases in NaCl concentration gave fractions eluting between 0.05M NaCl and 0.15M NaCl which were pooled and dialyzed against PBS. The solution was sterilized by passage through a 0.2 micron filter to yield a fluorescein labeled antibody.

Workup

  1. customafter which the solution was dialyzed against 0.015M NaCl, 0.01M sodium phosphate
  2. temperatureStep-wise increases in NaCl
  3. concentrationconcentration
  4. customgave fractions
  5. washeluting between 0.05M NaCl and 0.15M NaCl which
  6. customdialyzed against PBS
  7. filtrationfilter

Sources & evidence

ORD record
ord-eab8b51cbafc4314b3c58eebfc7b3740
Source dataset
uspto-grants-1984_12
Patent number
US04490473

Reaction notes

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