Galectin 3 Protein (LGALS3)
(1 validation)-
- 抗原 See all Galectin 3 (LGALS3) 蛋白
- Galectin 3 (LGALS3)
- 蛋白类型
- Recombinant
- 生物活性
- Active
-
宿主
- 人
-
资源
- 大肠杆菌（E. Coli）
- 应用范围
- Functional Studies (Func)
- 产品特性
- Recombinant Human Galectin-3 (LGALS3)
- 纯度
- >95 % as determined by SDS-PAGE
- 内毒素水平
- < 1.0 EU of endotoxin/μg of protein
-
Galectin 3 (LGALS3) (AA 2-250) protein (His tag)
LGALS3 宿主: 人 宿主: HEK-293 Cells Recombinant > 95 % as determined by Tris-Bis PAGE
Galectin 3 (LGALS3) (AA 2-264) protein (His tag)LGALS3 宿主: 小鼠 宿主: HEK-293 Cells Recombinant > 95 % as determined by Tris-Bis PAGE
Galectin 3 (LGALS3) (AA 2-262) protein (His tag)LGALS3 宿主: 大鼠 宿主: 大肠杆菌（E. Coli） Recombinant Greater than 95 % by SDS-PAGE gel analyses WB, ELISA
Galectin 3 (LGALS3) (AA 4-262) protein (His tag)LGALS3 宿主: Pig 宿主: 大肠杆菌（E. Coli） Recombinant Greater than 95 % by SDS-PAGE gel analyses WB, ELISA
Galectin 3 (LGALS3) (AA 1-264) protein (His tag)LGALS3 宿主: 小鼠 宿主: 大肠杆菌（E. Coli） Recombinant Greater than 95 % as determined by SDS-PAGE WB, ELISA
Galectin 3 (LGALS3) (AA 2-250) protein (His tag)LGALS3 宿主: 人 宿主: 大肠杆菌（E. Coli） Recombinant Greater than 95 % by SDS-PAGE gel analyses WB, ELISA
Galectin 3 (LGALS3) (AA 1-250) protein (GST tag)LGALS3 宿主: 人 宿主: 小麦胚 Recombinant WB, ELISA, AP, AA
-
- 应用备注
- Optimal working dilution should be determined by the investigator.
- 限制
- 仅限研究用
-
- by
- Sanja Dabelic, Ph.D., Associate Professor, Department of Biochemistry and Molecular Biology, Faculty of Pharmacy and Biochemistry, University of Zagreb
- No.
- #103788
- 日期
- 2019.06.17
- 抗原
- LGALS3
- Lot Number
- 158844928901
- Method validated
- Haemagglutination
- Positive Control
human erythrocytes (blood group A)
serum from uncompatible donor (blood group B)
- Negative Control
PBS
0.5mM lactose
- Notes
Passed. The recombinant human galectin-3 ABIN5509728 possesses lectin activity.
- Primary Antibody
- Secondary Antibody
- Full Protocol
- Isolate human erythrocytes and prepare trypsinized,glutaraldehyde-fixed erythrocytes
- Take blood by venipuncture from healthy volunteer (blood type A) in one lavender vacutainer tube (containing EDTA as anticoagulant) and centrifuged at 500×g for 5min at 4°C.
- Remove plasma and white cell ghost layer at the top of the pellet.
- Resuspend red blood cells in 10ml cold PBS by gently aspirating and expelling them with a pipette. Centrifuge red blood cells at 500×g for 5min at 4°C. Remove the supernatant and repeat 3x.
- Resuspend the pellet in 20ml PBS (Gibco) containing 0.05% trypsin and incubate at 37°C for 1h with an occasional shake.
- Centrifuge at 500xg for 5min at 4°C.
- Remove supernatant and resuspend erythrocytes in 10ml cold PBS.
- Wash pellet 3x as described above.
- Resuspend erythrocytes in 25ml PBS containing 1% glutaraldehyde and incubate for 1h at RT with continuous gentle shaking.
- Centrifuge at 500xg for 5min at 4°C.
- Remove supernatant and wash erythrocytes 2x with 0.1M glycine in PBS.
- Wash erythrocytes with PBS.
- Store erythrocytes as 4% (v/v) erythrocyte suspension at 4 °C until use (within one month of preparation).
- Isolation of human serum (blood type B)
- Take blood by venipuncture from healthy volunteer (blood type B) in one red topped vacutainer tube (additive-free vacutainer).
- Allow the blood to clot by leaving it undisturbed at room temperature for 30min.
- Centrifuge at 1000xg for 10min at 4°C.
- Remove serum supernatant and store at -20°C until use.
- Preparation of galectin-3 and lactose
- Reconstitute lyophilized galectin-3 (antibodies-online, ABIN5509728, lot 0158844928901) in sterile, 0.2μm filtered solution of 0.2% bovine serum albumin (BSA) in PBS, according to the manufacturers’ recommendations, to 100μg/ml.
- Dilute D-lactose in PBS to 2mM.
- Measurement of Hemagglutination
- Prepare samples in triplicates as indicated in Figure A in a total volume of 100µl in a microtiter V plate.
- Incubate plate for 1h and 24h at 4°C.
- Determine results by visual examination. Image acquisition on an Amersham Imager 600, Image capturing method colorimetric-transillumination.
- Experimental Notes
Galectin-3 is approximately 30kDa and, like all galectins, contains a carbohydrate-recognition-binding domain (CRD) of about 130 amino acids that enable the specific binding of β-galactosides. For several decades, hemagglutination assay has been used to obtain semi-quantitative data on the sugar binding and specificity of a particular lectin. Active lectins agglutinate erythrocytes by recognizing carbohydrates on the cell surface and forming a cross-linked network in suspension.
By serially diluting the lectin in a 96-well microtiter plate and adding a constant quantity of erythrocytes, the lectin activity can be estimated. Galectin-3 lectin activity is abolished by the addition of lactose, which binds to its CRD domain and consequently inhibits hemagglutination. The assay is usually performed on trypsinized, glutaraldehyde-fixed red blood cells because they are more sensitive to hemagglutination reactions. Addition of bovine serum albumin (BSA) to all wells prevents non-specific interactions and consequently non-specific auto-hemagglutination. As positive control, serum of a non-compatible blood type can be used. Negative control is PBS. Control for abolishing galectin-3 activity is lactose.
The hemagglutination assay is carried out by incubating human erythrocytes (blood group A or B) in the presence of increasing concentrations of galectin-3 in a microtiter V plate for 1h and 24h at RT in a final volume of 100µl. Galectin-3 recognizes glycans on both type A and type B red blood cells at submicromolar concentrations, according to the literature data, but does not bind to blood group O(H) at these concentrations. Commercially available 4% solutions of trypsinized, glutaraldehyde-fixed erythrocytes from various species and (e.g. rabbit) can also been used.
Pre-incubation of galectin-3 for 30min in the presence of 500µM lactose should cause prevention (inhibition) of hemagglutination.
Hemagglutination was detected in wells that contained the positive control without (wells A4-A6) and with lactose (wells A10-12), and in wells with galectin-3 at final concentrations 25μg/mL, 12.5μg/mL, and 5μg/mL (wells B1-9). A galectin-3 concentration of 1μg/ml was insufficient to cause complete hemagglutination (wells B10 to B12). Partial inhibition was detected in wells containing galectin-3 at 25µg/ml in the presence of 0.5mM lactose (well C1-3). At galectin-3 concentrations 12.5µg/ml, 5µg/ml, and 1µg/ml hemagglutination is completely abolished in the presence of 0.5mM lactose (well C4-12).
References:
https://www.ncbi.nlm.nih.gov/pubmed/1259723
https://www.ncbi.nlm.nih.gov/pubmed/10722666
https://www.ncbi.nlm.nih.gov/pubmed/22497898
https://www.ncbi.nlm.nih.gov/pubmed/27328612
https://www.ncbi.nlm.nih.gov/pubmed/30797745
https://www.ncbi.nlm.nih.gov/pubmed/18216021
生效 #103788 (Haemagglutination)!['独立验证'标志]( "成功验证")
!['独立验证'标志]( "成功验证")
Validation Images![Hemagglutination assay using recombinant human galectin-3 ABIN5509728. A. Plate Layout. B. and C. Pictures captured after 1h and 24h respectively.]()
Hemagglutination assay using recombinant human galectin-3 ABIN5509728. A. Plate Layout. B. and C. Pictures captured after 1h and 24h respectively.
Full Methods -
- 状态
- Lyophilized
- 溶解方式
- A quick spin of the vial followed by reconstitution in sterile distilled water to a concentration not less than 0.1 mg/mL is recommended. Please note, filter sterilization is a must following reconstitution. This solution can then be diluted into other buffers.
- 储存条件
- 4 °C,-20 °C
- 储存方法
- The lyophilized protein is stable for at least one year from date of receipt at -70°C. Upon reconstitution, this cytokine can be stored in working aliquots at 2° - 8°C for one month, or at -20°C for six months, with a carrier protein without detectable loss of activity. Avoid repeated freeze/thaw cycles.
-
- 抗原
- Galectin 3 (LGALS3)
- 别名
- Galectin-3 (LGALS3) (LGALS3 产品)
- 别名
- CBP 35 Protein, Gal-3 Protein, Galectin-3 Protein, leg3 Protein, galactoside-binding Protein, galectin-3 Protein, lectin Protein, soluble Protein, CBP35 Protein, GAL3 Protein, GALBP Protein, GALIG Protein, L31 Protein, LGALS2 Protein, MAC2 Protein, MGC52947 Protein, MGC75803 Protein, LGALS3 Protein, gal-3 Protein, GBP Protein, L-34 Protein, Mac-2 Protein, gal3 Protein, galectin 3 Protein, Galectin-3 Protein, avian beta-defensin 3 Protein, lectin, galactoside-binding, soluble, 3 L homeolog Protein, lectin, galactoside-binding, soluble, 3 Protein, galectin 2 Protein, lectin, galactose binding, soluble 3 Protein, LGALS3 Protein, leg3 Protein, AvBD3 Protein, lgals3.L Protein, lgals3 Protein, LGALS2 Protein, Lgals3 Protein
- 物质类
- Chemical
- 分子量
- 26,1
- 基因ID
- 3958
- UniProt
- P17931
- 途径
- RTK signaling
-
