Preparation method of breast cancer sentinel lymph node tracer agent indocyanine green-rituximab

A technology of rituximab and sentinel lymph node, which is applied in the field of preparation of breast cancer sentinel lymph node tracer indocyanine green-rituximab, can solve the problem of limited popularization, radionuclide pollution, and no detailed operation Steps and specific equipment dosage and other issues

Active Publication Date: 2015-07-01
王永胜
View PDF2 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, the dye method (methylene blue, patent blue and nano-carbon) locates the sentinel lymph node mainly through the preoperative dye around the primary breast tumor and (or) subcutaneously injecting the dye, and then following the dye-stained lymphatic vessels in the axillary area for fine dissection Find the first stained lymph node to determine the specific location of the sentinel lymph node. However, this method requires superb surgical anatomy skills, and there is still a high false-negative rate of sentinel lymph node biopsy after the learning curve, which leads to sentinel lymph node biopsy. The accuracy of the method is difficult to grasp; while the nuclide method ( 99m Tc-sulfur colloid) is used to locate the sentinel lymph node by preoperatively injecting radionuclide into the glands around the primary breast tumor and (or) subcutaneously, and then use the gamma detector to detect the radioactive count during the operation, and find the lymph node with a high count according to the radioactive count , so as to determine the specific location of the sentinel lymph node (lymph node greater than 10% of the highest radioactive count), although this method of sentinel lymph node biopsy has high accuracy and good specificity, but there are risks of radionuclide contamination and radiation damage to medical staff during actual operation problems, and the sulfur colloid has not been approved by the China Food and Drug Administration, which limits its promotion and use in clinical practice; the method of locating sentinel lymph nodes by fluorescence method (indocyanine green) is similar to the dye method, mainly After the preoperative injection of a fluorescent tracer around the primary breast tumor and/or subcutaneously, the first lymph node that is positive for fluorescence imaging is found by using a near-infrared fluorescence imaging device to follow the bright lymph vessels of fluorescence imaging during surgery , so as to locate the specific location of the sentinel lymph node. However, since ...

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Preparation method of breast cancer sentinel lymph node tracer agent indocyanine green-rituximab
  • Preparation method of breast cancer sentinel lymph node tracer agent indocyanine green-rituximab
  • Preparation method of breast cancer sentinel lymph node tracer agent indocyanine green-rituximab

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] The preparation method of breast cancer sentinel lymph node tracer indocyanine green-rituximab comprises the following steps:

[0037] Under room temperature, according to the principles of aseptic operation, 100 mg of rituximab (Rituximab 100 mg, American Roche Company) was prepared into 10 mg / ml with 10 ml of sterilized water for injection (5 ml of sterilized water for injection, Tianjin Pharmaceutical Group Xinzheng Co., Ltd.) 25mg of indocyanine green (indocyanine green for injection 25mg Dandong Yichuang Pharmaceutical Co., Ltd.) was prepared with 10ml sterile water for injection to a concentration of 2.5mg / ml solution, put it in a 50ml sterile centrifuge tube and keep it away from light for later use. At room temperature, take 6 parts of 0.5ml rituximab solution that has been prepared and place them in 10ml centrifuge tubes, and then take rituximab and indocyanine green with a mass ratio of 3 :1, 4:1, 6:1, 12:1, 30:1 and 32:1 of the corresponding volume of 2.5mg / ...

Embodiment 2

[0038] Embodiment 2 detects the molecular integrity of indocyanine green-rituximab coupling reaction product:

[0039] Use 8% non-reducing SDS polyacrylamide gel electrophoresis to detect the molecular integrity of indocyanine green-rituximab, and use rituximab as a positive control to determine the indocyanine green-rituximab Integrity of the molecule, the mass ratio of rituximab to indocyanine green in Example 1 was 4:1, 6:1, 12:1, 30:1 and 32:1 The coupling reactant did not appear Obvious degradation bands, such as Figure 4 As shown (the results of the gel spectrum of the coupling reactants in the above mass ratios are consistent, and no degradation bands appear), indicating that the molecular structures of the coupling products of the two are complete.

Embodiment 3

[0040] Embodiment 3 detects the immune activity of indocyanine green-rituximab conjugate:

[0041] Double-antibody sandwich indirect enzyme-linked immunoassay (ELISA) method to detect the immune activity of indocyanine green-rituximab, rabbit anti-mouse IgG-Fab antibody is the coating antibody, and the mass fraction is 1% bovine serum albumin blocking For 1 hour, add the conjugates whose mass ratios of rituximab and indocyanine green in Example 1 are 4:1, 6:1, 12:1, 30:1 and 32:1, and avoid Light reaction for 2 hours, adding horseradish peroxidase-labeled goat anti-human IgG-Fc fragment antibody, reacting in the dark for 1 hour at room temperature, developing color with o-phenylenediamine hydrochloride, and measuring A 412nm value, such as Figure 5 As shown, the results show that the mass ratio of rituximab to indocyanine green is 4:1, 6:1, 12:1, 30:1 and 32:1, and the coupling reaction is positive, OD 412 No significant difference was found, which indicates that the immuno...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

PropertyMeasurementUnit
Diameteraaaaaaaaaa
Molecular weightaaaaaaaaaa
Login to view more

Abstract

The invention discloses a preparation method of breast cancer sentinel lymph node tracer agent indocyanine green-rituximab. The method comprises the following steps: slowly dropping an indocyanine green solution into a rituximab solution, rapidly oscillating, mixing evenly and reacting; carrying out coupled reaction, carrying out centrifugal precipitation on a centrifuge tube with a coupled reactant, and observing presence and absence of sediments; if the sediments are generated, removing sediments at the bottom of the centrifuge tube, taking supernate and putting the supernate into a dialysis bag made of a semi-permeable membrane; immersing the dialysis bag into dialysate for sterilizing injection water to dialyze; and bonding the indocyanine green on rituximab through coupled reaction, so as to prepare a novel indocyanine green-rituximab combined tracer agent. The preparation method is simple, safe, and free of radiation; the prepared indocyanine green-rituximab can meet the requirements of developing and positioning of the sentinel lymph node.

Description

technical field [0001] The invention relates to a method for preparing a breast cancer sentinel lymph node specific tracer indocyanine green-rituximab. Background technique [0002] The sentinel lymph node is the first lymph node of breast cancer metastasis, and its pathological conditions have important guiding value for breast cancer surgery and postoperative comprehensive treatment. Sentinel lymph node biopsy is a technical method for lymph node staging of breast cancer. Accurate positioning of sentinel lymph nodes is a technical field that has been concerned in research on sentinel lymph node biopsy of breast cancer. Traditional tracers for sentinel lymph node localization include dyes, nuclides, and fluorescence, all of which are non-specific tracers. Among them, the dye method (methylene blue, patent blue and nano-carbon) locates the sentinel lymph node mainly through the preoperative dye around the primary breast tumor and (or) subcutaneously injecting the dye, and t...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): A61K49/00
Inventor 王永胜丛斌斌孙晓宋现让杨国仁曹晓珊田崇麟张燕李盼盼
Owner 王永胜
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products