nSMOL Antibody BA Kit

nSMOL (nano-surface and molecular orientation limited proteolysis) is Shimadzu’s proprietary, innovative technique that enables selective proteolysis of the Fab region of monoclonal antibodies. The nSMOL Antibody BA Kit is a ready-to-use reagent kit for collecting monoclonal antibodies from blood or other biological samples using immunoglobulin collection resin, and then performing selective proteolysis of the Fab region of these antibodies via trypsin-immobilized nanoparticles. Variable region-derived peptides produced by limited proteolysis can then be quantified via MRM measurements utilizing a high-performance LCMS-8050/8060 triple quadrupole liquid chromatograph mass spectrometer. An unparalleled convenient and rapid workflow provided by the nSMOL Antibody BA Kit dramatically improves the productivity and robustness of LCMS mAb bioanalysis.

  • Faster, Less Expensive Method Development
    Dramatically improves response and quantitative repeatability. No capture antibodies or ligands required.

  • nSMOL Proteolysis
    Selective collection of Fab peptides. Limits contamination from excessive peptides or trypsin.

  • Performance
    Highly sensitive and accurate assays are possible for a variety of antibodies.

  • Highly Versatile
    Applicable to a wide variety of pharmaceutical antibodies.

 

Simple Workflow

This kit enables highly reproducible data and avoids the troublesome steps of denaturing, reduction, and alkyation normally associated with protein digestion. There is also no need for solid phase extraction after reaction. After nSMOL preparation, samples can be injected directly onto the LCMS.

Kit Contents

The nSMOL Antibody BA Kit provides prepared reagents and protocols for sample prep via the nSMOL method.

Product Specifications & Ordering Information

Item Specifications
P/N 225-32250-91
Detection Targets Monoclonal antibodies in serum or plasma (including antibody pharmaceuticals)
Number of Measurement 100 assays
Approximate volume of sample 90 μl (before LCMS injection)
Storage Temperature Specified for each reagent
Expiration Date 18 months from the date of shipment if unopened and stored at 4℃
(*For FG BeadsTM Trypsin DARTTM, 18 months from the date of shipment if stored at -80℃)

Product Configuration

Reagent Quantity Capacity Storage
Temperature
Immunoglobulin Collection Resin 2 1.3 mL/each 4℃
Wash(Binding)Solution 1 1 80 ml 4℃
Wash Solution 2 1 80 ml 4℃
Reaction Solution 1 10 ml 4℃
Enhanced Reaction Solution 1 Freeze-dried 4℃
Reaction Stop Solution 1 1 ml 4℃
FG Beads™ Trypsin DART™ 1 1.1 ml -20℃*1

*1 If it is being stored for a month or longer, store it at -80 °C.

The following consumables are purchased separately.

Specied Consumables Manufacturer P/N
nSMOL reaction socket tube SHIMADZU 225-32260-91
225-32260-92
Ultrafree-MC, GV 0.22 μm Merck Millipore UFC30GV00
UFC30GVNB
Micro tube 2mL, PP SARSTEDT 72.708

Note: For other equipment required, please refer to instruction manual.

Recommended Items

Refer to the recommended specifications and reference products in the following table, and make preparations accordingly.

Required Items Recommended Specifications Reference Products
5 μm filtration filter Prepare a syringe and syringe filter with a pore size of 5 μm.
Millex-SV 5.0 μm, PVDF, 25 mm,
PVC housing, non-sterile,
SLSV025NB, Merck Millipore
0.8 μm filtration filter Prepare a syringe and syringe filter with a pore size of 0.8 μm. Millex-AA Syringe Filter Unit, 0.80 μm,
mixed cellulose esters, 25 mm, PVC housing non-sterile,
SLAA025NB, Merck Millipore
2 mL Sample Tube ・Polypropylene
・Round bottom tube
Item name:Safe-Lock Tubes
P/N:0030 120.094
Sold by:Eppendorf
Micro Tube Mixer ・Used at room temperature or 50 ℃.
・Can be installed inside the incubator.
・Set available with a 1.5 mL tube, a 2.0 mL tube, and a filter tube.
Item name:Micro tube mixer
Model:MT-400
Sold by:Tomy Seiko Co., Ltd
Incubator ・Incubator with a humidistat, or an incubator that a tray filled with water can be placed inside in order to maintain saturation vapor pressure
・Big enough that the above-mentioned micro tube mixer can be installed
Item name:Incubator with a freezer
Model:MIR-254-PJ
Sold by:Panasonic Healthcare Co., Ltd.
Micro Pipetter, Tip Tip without plasticizer to avoid plasticizer contamination in the MS (colorless) (1) Brand name:Rainin Sold by:Mettler Toledo
(2) Brand name:Biohit Sold by:Sartorius
Desktop Centrifuge Capable of centrifugation of 1.5 mL tubes, 2.0 mL tubes, and filter tubes at 10,000 μg and 12,000 μg Product series name:MX series high speed micro centrifuge
Sold by:Tomy Seiko Co., Ltd
Magnetic Tube Stand Capable of magnetically separating FG beads trypsin DART Item name:Magnet stand
Model:TA4899N12
Sold by:Tamagawa Seiki Co., Ltd.
LCMS Vials Low adsorption type, or vials without plasticizer Item name:TORAST-H Bio Vial  Item name:TORAST-H Bio Vial P/N 980-19983
Sold by:Shimadzu 
Analysis Column Capable of peptide separation and analysis

Item name:Shim-pack GISS
Inner dia. 2.1 mm ×length 50 mm,
particle size 1.9 μm
P/N: 227-30048-01
Sold by:Shimadzu 

Internal Standard Solution Capable of sensitivity and retention time correction Item name:ProteoMass P14R MALDI-MS standard
Catalog no:P2613
Sold by:Sigma-Aldrich
 

References / Scientific publications

  1. What sample preparation should be chosen for targeted MS monoclonal antibody quantification in human serum?
    Vialaret J, Broutin S, Pugnier C, Santelé S, Jaffuel A, Barnes A, Tiers L, Pelletier L, Lehmann S, Paci A, Hirtz C.
    Bioanalysis. 2018 May 1;10(10):723-735. doi: 10.4155/bio-2018-0027. Epub 2018 May 17.
    PMID: 29771137
     
  2. Structure-Indicated LC-MS/MS Bioanalysis of Therapeutic Antibodies.
    Iwamoto N, Shimada T.
    Methods Mol Biol. 2022;2313:187-205. doi: 10.1007/978-1-0716-1450-1_11.
    PMID: 34478139
     
  3. Iwamoto N, Shimada T.
    Pharmacol Ther. 2018 May;185:147-154. doi: 10.1016/j.pharmthera.2017.12.007. Epub 2017 Dec 22.
    PMID: 29274706 Review.
     
  4. Koguchi Y, Iwamoto N, Shimada T, Chang SC, Cha J, Curti BD, Urba WJ, Piening BD, Redmond WL.
    J Immunother Cancer. 2021 Oct;9(10):e002663. doi: 10.1136/jitc-2021-002663.
    PMID: 34620702 Free PMC article.
     
  5. Iwamoto N, Takanashi M, Shimada T, Sasaki J, Hamada A.
    AAPS J. 2019 Aug 20;21(6):101. doi: 10.1208/s12248-019-0369-z.
    PMID: 31432293
     
  6. Iwamoto N, Shimada T, Umino Y, Aoki C, Aoki Y, Sato TA, Hamada A, Nakagama H.
    Analyst. 2014 Feb 7;139(3):576-80. doi: 10.1039/c3an02104a.
    PMID: 24326404
     
  7. Iwamoto N, Shimomura A, Tamura K, Hamada A, Shimada T.
    J Pharm Biomed Anal. 2017 Oct 25;145:33-39. doi: 10.1016/j.jpba.2017.06.032. Epub 2017 Jun 19.
    PMID: 28648785
     
  8. Iwamoto N, Yokoyama K, Takanashi M, Yonezawa A, Matsubara K, Shimada T.
    Curr Pharm Biotechnol. 2018;19(6):495-505. doi: 10.2174/1389201019666180703093517.
    PMID: 29968534 Free PMC article.
     
  9. Iwamoto N, Shimada T, Terakado H, Hamada A.
    J Chromatogr B Analyt Technol Biomed Life Sci. 2016 Jun 15;1023-1024:9-16. doi: 10.1016/j.jchromb.2016.04.038. Epub 2016 Apr 27.
    PMID: 27155936
     
  10. Iwamoto N, Takanashi M, Yokoyama K, Yonezawa A, Denda M, Hashimoto M, Tanaka M, Ito H, Matsuura M, Yamamoto S, Honzawa Y, Matsubara K, Shimada T.
    J Immunol Methods. 2019 Sep;472:44-54. doi: 10.1016/j.jim.2019.06.014. Epub 2019 Jun 12.
    PMID: 31201793
     
  11. Iwamoto N, Yokoyama K, Takanashi M, Yonezawa A, Matsubara K, Shimada T.
    Pharmacol Res Perspect. 2018 Jul 24;6(4):e00422. doi: 10.1002/prp2.422. eCollection 2018 Jul.
    PMID: 30062014 Free PMC article.
     
  12. Iwamoto N, Umino Y, Aoki C, Yamane N, Hamada A, Shimada T.
    Drug Metab Pharmacokinet. 2016 Feb;31(1):46-50. doi: 10.1016/j.dmpk.2015.11.004. Epub 2015 Nov 30.
    PMID: 26830079
     
  13. Iwamoto N, Takanashi M, Umino Y, Aoki C, Hamada A, Shimada T.
    Bioanalysis. 2016 May;8(10):1009-20. doi: 10.4155/bio-2016-0018. Epub 2016 Mar 14.
    PMID: 26972866
     
  14. Iwamoto N, Takanashi M, Hamada A, Shimada T.
    Biol Pharm Bull. 2016 Jul 1;39(7):1187-94. doi: 10.1248/bpb.b16-00230. Epub 2016 Apr 28.
    PMID: 27150271
     
  15. Iwamoto N, Yonezawa A, Matsubara K, Shimada T.
    J Pharm Biomed Anal. 2019 Feb 5;164:467-474. doi: 10.1016/j.jpba.2018.11.019. Epub 2018 Nov 8.
    PMID: 30447535
     
  16. Iwamoto N, Shimada T.
    Drug Metab Pharmacokinet. 2019 Feb;34(1):19-24. doi: 10.1016/j.dmpk.2018.10.002. Epub 2018 Oct 19.
    PMID: 30392772 Review.
     
  17. Iwamoto N, Hamada A, Shimada T.
    Anal Biochem. 2018 Jan 1;540-541:30-37. doi: 10.1016/j.ab.2017.11.002. Epub 2017 Nov 9.
    PMID: 29128290

For Research Use Only. Not for use in diagnostic procedures.

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