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HyperSep Columns Application Notebook – Removing Uncertainty by Applying Science to SPE Product Literature

What are the substrates for the matrix metalloproteases (MMPs)? Product FAQ

Answer

The substrates for the matrix metalloproteases (MMPs) are:

*MMP-1: tissue collagenase: collagen 1, 2, 3, 4, 6, & 10

*MMP-2: gelatinase: gelatins, collagens 4, 5, & 7

*MMP-3: stromleysin-2: casein, fibronectin, laminin, elastin

*MMP-7: matrilysin: casein

*MMP-8: neutrophil collagenase: collagen

*MMP-9: type 4 collagenase: gelatin

*MMP-10: stromelysin-2: casein

*MMP-12: metalloelastase: elastin

Answer Id:: E10735

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What are the 10 CytoScan assay processing steps listed in order? Product FAQ

Answer

The 10 assay processing steps, as listed in the CytoScan Assay User Manual (Cat. No. 703038) are the following:

1. Genomic DNA Preparation
2. NSP1 Restriction Enzyme Digestion
3. Ligation
4. PCR
5. PCR Product Purification
6. Quantitation
7. Fragmentation
8. Labeling
9. Hybridization
10. Wash, Stain, Scanning

Answer Id:: E13962

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Pharmacokinetics and pharmacodynamics of LGD-3303 [9-chloro-2-ethyl-1-methyl-3-(2,2,2-trifluoroethyl)-3H-pyrrolo-[3,2-f]quinolin-7( 6H)-one], an orally available nonsteroidal-selective androgen receptor modulator. Citations & References

  • Authors: Vajda EG, Lopez FJ, Rix P, Hill R, Chen Y, Lee KJ, O'Brien Z, Chang WY, Meglasson MD, Lee YH
  • Journal: J Pharmacol Exp Ther
  • PubMed ID: 19017848

How can I cross-link antibodies to Dynabeads magnetic beads? Product FAQ

Answer

Here is a method for cross-linking of IgG immobilized Dynabeads Protein G magnetic beads (5 µg goat a-HSA immobilized to 50 µL Dynabeads Protein G magnetic beads, in PBST), with BS^3 linker from Thermo Scientific (Cat. No. 21580):

(1) BS^3 Stock and Conjugation solutions have to be freshly prepared just before use (i.e., make them after IgG immobilization is complete).
(2) Prepare 100 mM BS^3 in Conjugation Buffer (Stock Solution). Proceed to making a 5 mM solution by diluting in Conjugation Buffer.
(3) Wash the Ig-coupled Dynabeads Protein G magnetic beads twice in 200 µL Conjugation Buffer. Place on magnet and discard supernatant.
(4) Resuspend the Dynabeads magnetic beads in 250 µL 5 mM BS^3.
(5) Incubate for 30 min with tilting/rotation at room temperature.
(6) Quench the cross-linking reaction by adding 12.5 µL Quenching Buffer
(7) Incubate for 15 min with tilting/rotation at room temperature.
(8) Wash cross-linked Dynabeads magnetic beads three times with 200 µL PBST (or IP buffer of your choice), place on magnet, discard supernatant
(9) Proceed with your IP and antigen elution

BS^3 Conjugation buffer: 20 mM sodium phosphate, 0.15 M NaCl, pH 7-9

BS^3 Quenching buffer: 1 M Tris HCl, pH 7.5.

Answer Id:: E6019

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S15829EX SDS

Catalog #

I am interested in performing Luminex assays using BioSource kits, and I have a Luminex xMAP system. Besides the kits and system, what other reagents and equipment will I need? Product FAQ

Answer

The following is a list of general lab supplies that are required for running BioSource immunoassays on the Luminex xMAP system:
1) Vacuum manifold
2) Sonicating water bath
3) Orbital shaker
4) Vortexer
5) Repeating and/or multi-channel pipetter (not required, but recommended)
6) Calibrated adjustable precision pipettes, with disposable plastic tips
7) Glass/plastic tubes and racks for preparing reagents
8) Graduated cylinder and container for preparing wash solution
9) Aluminum foil
10) Deionized or distilled water.

Answer Id:: E5180

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What are the molecular weights of the proteins in the HiMark Unstained Protein Standard? Product FAQ

Answer

The HiMark Unstained Protein Standard allows the accurate estimation of molecular weight of high molecular weight proteins on NuPAGE Tris-Acetate Gels with Tris-Acetate SDS buffer system. Here are the molecular weights of the proteins in the HiMark Unstained Protein Standard:

- Band 1: 500 kDa
- Band 2: 290 kDa
- Band 3: 240 kDa
- Band 4: 160 kDa
- Band 5: 116 kDa
- Band 6: 97 kDa
- Band 7: 66 kDa
- Band 8: 55 kDa
- Band 9: 40 kDa

Answer Id:: E11691

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What are the default or standard values for the Procise System pressure regulators? Product FAQ

Answer

Reg. 1 (R1) = 2.5 psi
Reg. 2 (R2) = 1.0 psi
Reg. 3 (R3) = 1.5 psi , set to 0.8 psi for gas-phase sequencing
Reg. 4 (S1, S2 and S3) = 1.7 psi
Reg. 5 (Cart Dry) = 3.5 psi
Reg. 6 (R4 and S4) = 3.5 psi
Reg. 7 (R5, X1 and X2) = 2.5 psi
Reg. 8 (X3 and Flask Dry) = 3.0 psi
Reg. 9 (Flask Bubble) = 1.8 psi
Reg. 10 (Load Injector) = 1.0 psi

Answer Id:: E1893

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How can I increase the sensitivity of my Northern hybridizations? Product FAQ

Answer

Please see below the top ten ways to increase sensitivity of your Northern hybridizations:

1) Increase the amount of RNA loaded in each lane (up to 30 mg).
2) Use poly(A) RNA instead of total RNA; 10 mg of poly(A) RNA is ~300-350 mg total RNA (3-5%).
3) Switch to ULTRAhyb Ultrasensitive Hybridization Buffer.
4) Switch from DNA to RNA probes.
5) Use downward alkaline capillary transfer.
6) Use an optimal hybridization temperature.
7) Use a freshly synthesized probe.
8) Use a high specific activity probe (10^8 to 10^9 cpm/mg).
9) Increase exposure time (it can take up to 3 days to see low-abundance messages with radiolabeled probes).
10) Follow the manufacturer's recommendations to crosslink the RNA to the membrane.

Read more about these suggestions here (https://www.thermofisher.com/us/en/home/references/Invitrogen-tech-support/northern-analysis/general-articles/ten-ways-to-increase-the-sensitivity-of-northern-hybridizations.html).

Answer Id:: E8031

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What is the formulation of your TAP media? Product FAQ

Answer

We offer premade TAP growth media optimized for Chlamydomonas (Cat. No. A1379801). Please see the formulation below:

TAP Media
EDTA (Disodium Salt), EDTA, 0.005%, CAS: 60-00-4
Zinc Sulphate Heptahydrate, ZnSO4-7H2O, 0.002%, CAS: 7446-20-0
Boric Acid, H3BO3, 0.001%, CAS: 10043-35-3
Manganese Chloride Tetrahydrate, MnCl2-4H2O, 0.005%, CAS: 1/5/7773
Cobalt Chloride Hexahydrate, CoCl2-6H2O, 0.002%, CAS: 7791-13-1
Copper Sulphate Pentahydrate, CuSO4·5H2O, 0.002%, CAS: 7758-98-7
Ammonium heptamolybdate, (NH4)6Mo7O24·4H2O, 0.001%, CAS:12027-67-7
Iron(II) Sulphate, FeSO4·7H2O, 0.005%, CAS:7720-78-7
Potassium Phosphate Dibasic, K2HPO4, 0.011%, CAS:11/4/7758
Potassium dihydrogen phosphate, KH2PO4, 0.005%, CAS:7778-77-0
Ammonium chloride, NH4Cl, 0.038%, CAS:12125-02-9
Magnesium Sulfate Heptahydrate, MgSO4 . 7H2O, 0.010%, CAS:10034-99-8
Calcium Chloride Dihydrate, CaCl2 . 2H2O, 0.005%, CAS:10035-04-8
Tris, (HOCH2)3CNH2, 0.242%, CAS:77-86-1
Glacial Acetic Acid, CH3COOH, 0.11%, CAS:64-19-7

Answer Id:: E9569

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Custom Fluid Transfer Service Product Literature

How can I isolate mouse spleen cells? Product FAQ

Answer

To isolate mouse spleen cells:
(1) Remove the spleen from the animal as fast as possible.
(2) Transfer the spleen to a 50 mL tube containing 20 mL cold Dulbecco's PBS with 0.1% BSA, 2 mM EDTA (buffer 1).
(3) Use sterile scissors to split the spleen in two.
(4) Transfer the two halves to a cell strainer (filter with large pores) premoistened with buffer 1. Place the cell strainer over a new empty 50 mL tube.
(5) Using the piston (the part with the rubber) from a 2 mL syringe, squeeze the spleen cells out, flushing them through the cell strainer with a total of 50 mL buffer 1.
(6) Centrifuge the mixture at 300 x g for 10 min at 4 degrees C.
(7) Resuspend the cell pellet in 5 mL of buffer 1 and filter through a new cell strainer.
(8) Add more buffer 1 and centrifuge at 300 x g for 10 min at 4 degrees C.
(9) Resuspend in buffer 1.

Answer Id:: E6144

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How can I crosslink antibodies to Dynabeads Protein G magnetic beads? Product FAQ

Answer

Here is a method for crosslinking of IgG immobilized Dynabeads Protein G magnetic beads (5 µg goat a-HSA immobilized to 50 µL Dynabeads Protein G magnetic beads, in PBST), with BS3 linker (Cat. No. 21580):

1. BS3 Stock and Conjugation solutions have to be freshly prepared just before use (i.e., make them after IgG immobilization is complete).
2. Prepare 100 mM BS3 in Conjugation Buffer (Stock Solution). Proceed to making a 5 mM solution by diluting in Conjugation Buffer.
3. Wash the Ig-coupled Dynabeads Protein G magnetic beads twice in 200 µL Conjugation Buffer. Place on magnet and discard supernatant.
4. Resuspend the Dynabeads magnetic beads in 250 µL 5 mM BS3.
5. Incubate for 30 min with tilting/rotation at room temperature.
6. Quench the crosslinking reaction by adding 12.5 µL Quenching Buffer
7. Incubate for 15 min with tilting/rotation at room temperature.
8. Wash crosslinked Dynabeads magnetic beads three times with 200 µL PBST (or IP buffer of your choice), place on magnet, discard supernatant 9. Proceed with your IP and antigen elution

BS3 Conjugation buffer: 20 mM sodium phosphate, 0.15 M NaCl, pH 7-9.
BS3 Quenching buffer: 1 M Tris HCl, pH 7.5.

Answer Id:: E13021

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