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Pulse–Chase Analysis for Studies of MHC Class II Biosynthesis, Maturation, and Peptide Loading

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Book cover Antigen Processing

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 960))

Abstract

Pulse–chase analysis is a commonly used technique for studying the synthesis, processing and transport of proteins. Cultured cells expressing proteins of interest are allowed to take up radioactively labeled amino acids for a brief interval (“pulse”), during which all newly synthesized proteins incorporate the label. The cells are then returned to nonradioactive culture medium for various times (“chase”), during which proteins may undergo conformational changes, trafficking, or degradation. Proteins of interest are isolated (usually by immunoprecipitation) and resolved by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and the fate of radiolabeled molecules is examined by autoradiography. This chapter describes a pulse–chase protocol suitable for studies of major histocompatibility complex (MHC) class II biosynthesis and maturation. We discuss how results are affected by the recognition by certain anti-class II antibodies of distinct class II conformations associated with particular biosynthetic states. Our protocol can be adapted to follow the fate of many other endogenously synthesized proteins, including viral or transfected gene products, in cultured cells.

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Abbreviations

MHC:

Major histocompatibility complex

ER:

Endoplasmic reticulum

Abs:

Antibodies

Ii:

Invariant chain

B-LCL:

EBV-transformed B-lymphoblastoid cell lines

SDS-PAGE:

Sodium dodecyl sulfate polyacrylamide gel electrophoresis

MIIC:

MHC class II compartments

LIP:

Leupeptin-induced polypeptides

CLIP:

Class-II-associated invariant chain peptides

Cys/Met:

Cysteine/methionine

PAS:

Protein A Sepharose

PGS:

Protein G Sepharose

IP:

Immunoprecipitation

APCs:

Antigen presenting cells

SLB:

Laemmli sample loading buffer

PMSF:

Phenylmethylsulfonyl fluoride

References

  1. Jamieson JD, Palade GE (1967) Intracellular transport of secretory proteins in the pancreatic exocrine cell. I. Role of the peripheral elements of the Golgi complex. J Cell Biol 34(2):577–596

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Jamieson JD, Palade GE (1967) Intracellular transport of secretory proteins in the pancreatic exocrine cell. II. Transport to condensing vacuoles and zymogen granules. J Cell Biol 34(2):597–615

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. De Riva A, Deery MJ, McDonald S, Lund T, Busch R (2010) Measurement of protein synthesis using heavy water labeling and peptide mass spectrometry: discrimination between major histocompatibility complex allotypes. Anal Biochem 403(1–2):1–12

    Article  PubMed  PubMed Central  Google Scholar 

  4. Jones PP, Murphy DB, Hewgill D, McDevitt HO (1979) Detection of a common polypeptide chain in I-A and I-E sub-region immunoprecipitates. Mol Immunol 16(1):51–60

    Article  PubMed  Google Scholar 

  5. Machamer CE, Cresswell P (1982) Biosynthesis and glycosylation of the invariant chain associated with HLA-DR antigens. J Immunol 129(6):2564–2569

    CAS  PubMed  Google Scholar 

  6. Busch R, Cloutier I, Sekaly RP, Hämmerling GJ (1996) Invariant chain protects class II histocompatibility antigens from binding intact polypeptides in the endoplasmic reticulum. EMBO J 15(2):418–428

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Bakke O, Dobberstein B (1990) MHC class II-associated invariant chain contains a sorting signal for endosomal compartments. Cell 63(4):707–716

    Article  CAS  PubMed  Google Scholar 

  8. West MA, Lucocq JM, Watts C (1994) Antigen processing and class II MHC peptide-loading compartments in human B-lymphoblastoid cells. Nature 369(6476):147–151

    Article  CAS  PubMed  Google Scholar 

  9. Tulp A, Verwoerd D, Dobberstein B, Ploegh HL, Pieters J (1994) Isolation and characterization of the intracellular MHC class II compartment. Nature 369(6476):120–126

    Article  CAS  PubMed  Google Scholar 

  10. Qiu Y, Xu X, Wandinger-Ness A, Dalke DP, Pierce SK (1994) Separation of subcellular compartments containing distinct functional forms of MHC class II. J Cell Biol 125(3):595–605

    Article  CAS  PubMed  Google Scholar 

  11. Peters PJ, Neefjes JJ, Oorschot V, Ploegh HL, Geuze HJ (1991) Segregation of MHC class II molecules from MHC class I molecules in the Golgi complex for transport to lysosomal compartments. Nature 349(6311):669–676

    Article  CAS  PubMed  Google Scholar 

  12. Blum JS, Cresswell P (1988) Role for intracellular proteases in the processing and transport of class II HLA antigens. Proc Natl Acad Sci USA 85(11):3975–3979

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Maric MA, Taylor MD, Blum JS (1994) Endosomal aspartic proteinases are required for invariant-chain processing. Proc Natl Acad Sci USA 91(6):2171–2175

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Riese RJ, Wolf PR, Bromme D, Natkin LR, Villadangos JA, Ploegh HL, Chapman HA (1996) Essential role for cathepsin S in MHC class II-associated invariant chain processing and peptide loading. Immunity 4(4):357–366

    Article  CAS  PubMed  Google Scholar 

  15. Neefjes JJ, Ploegh HL (1992) Inhibition of endosomal proteolytic activity by leupeptin blocks surface expression of MHC class II molecules and their conversion to SDS resistance alpha beta heterodimers in endosomes. EMBO J 11(2):411–416

    CAS  PubMed  PubMed Central  Google Scholar 

  16. Riberdy JM, Newcomb JR, Surman MJ, Barbosa JA, Cresswell P (1992) HLA-DR molecules from an antigen-processing mutant cell line are associated with invariant chain peptides. Nature 360(6403):474–477

    Article  CAS  PubMed  Google Scholar 

  17. Sette A, Ceman S, Kubo RT, Sakaguchi K, Appella E, Hunt DF, Davis TA, Michel H, Shabanowitz J, Rudersdorf R et al (1992) Invariant chain peptides in most HLA-DR molecules of an antigen-processing mutant. Science 258(5089):1801–1804

    Article  CAS  PubMed  Google Scholar 

  18. Denzin LK, Cresswell P (1995) HLA-DM induces CLIP dissociation from MHC class II alpha beta dimers and facilitates peptide loading. Cell 82(1):155–165

    Article  CAS  PubMed  Google Scholar 

  19. Sherman MA, Weber DA, Jensen PE (1995) DM enhances peptide binding to class II MHC by release of invariant chain-derived peptide. Immunity 3(2):197–205

    Article  CAS  PubMed  Google Scholar 

  20. Sloan VS, Cameron P, Porter G, Gammon M, Amaya M, Mellins E, Zaller DM (1995) Mediation by HLA-DM of dissociation of peptides from HLA-DR. Nature 375(6534):802–806

    Article  CAS  PubMed  Google Scholar 

  21. Lovitch SB, Pu Z, Unanue ER (2006) Amino-terminal flanking residues determine the conformation of a peptide-class II MHC complex. J Immunol 176(5):2958–2968

    Article  CAS  PubMed  Google Scholar 

  22. Germain RN, Rinker AG Jr (1993) Peptide binding inhibits protein aggregation of invariant-chain free class II dimers and promotes surface expression of occupied molecules. Nature 363(6431):725–728

    Article  CAS  PubMed  Google Scholar 

  23. Bikoff EK, Huang LY, Episkopou V, van Meerwijk J, Germain RN, Robertson EJ (1993) Defective major histocompatibility complex class II assembly, transport, peptide acquisition, and CD4+ T cell selection in mice lacking invariant chain expression. J Exp Med 177(6):1699–1712

    Article  CAS  PubMed  Google Scholar 

  24. Viville S, Neefjes J, Lotteau V, Dierich A, Lemeur M, Ploegh H, Benoist C, Mathis D (1993) Mice lacking the MHC class II-associated invariant chain. Cell 72(4):635–648

    Article  CAS  PubMed  Google Scholar 

  25. Busch R, Rinderknecht CH, Roh S, Lee AW, Harding JJ, Burster T, Hornell TM, Mellins ED (2005) Achieving stability through editing and chaperoning: regulation of MHC class II peptide binding and expression. Immunol Rev 207:242–260

    Article  CAS  PubMed  Google Scholar 

  26. Rinderknecht CH, Roh S, Pashine A, Belmares MP, Patil NS, Lu N, Truong P, Hou T, Macaubas C, Yoon T, Wang N, Busch R, Mellins ED (2010) DM influences the abundance of major histocompatibility complex class II alleles with low affinity for class II-associated invariant chain peptides via multiple mechanisms. Immunology 131(1):18–32

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Busch R, Doebele RC, von Scheven E, Fahrni J, Mellins ED (1998) Aberrant intermolecular disulfide bonding in a mutant HLA-DM molecule: implications for assembly, maturation, and function. J Immunol 160(2):734–743

    CAS  PubMed  Google Scholar 

  28. Pashine A, Busch R, Belmares MP, Munning JN, Doebele RC, Buckingham M, Nolan GP, Mellins ED (2003) Interaction of HLA-DR with an acidic face of HLA-DM disrupts sequence-dependent interactions with peptides. Immunity 19(2):183–192

    Article  CAS  PubMed  Google Scholar 

  29. Neefjes JJ, Hämmerling GJ, Momburg F (1993) Folding and assembly of major histocompatibility complex class I heterodimers in the endoplasmic reticulum of intact cells precedes the binding of peptide. J Exp Med 178(6):1971–1980

    Article  CAS  PubMed  Google Scholar 

  30. Nijenhuis M, Neefjes J (1994) Early events in the assembly of major histocompatibility complex class II heterotrimers from their free subunits. Eur J Immunol 24(1):247–256

    Article  CAS  PubMed  Google Scholar 

  31. Dornmair K, Rothenhäusler B, McConnell HM (1989) Structural intermediates in the reactions of antigenic peptides with MHC molecules. Cold Spring Harb Symp Quant Biol 54(Pt 1):409–416

    Article  CAS  PubMed  Google Scholar 

  32. Miyazaki T, Wolf P, Tourne S, Waltzinger C, Dierich A, Barois N, Ploegh H, Benoist C, Mathis D (1996) Mice lacking H2-M complexes, enigmatic elements of the MHC class II peptide-loading pathway. Cell 84(4):531–541

    Article  CAS  PubMed  Google Scholar 

  33. Roche PA, Marks MS, Cresswell P (1991) Formation of a nine-subunit complex by HLA class II glycoproteins and the invariant chain. Nature 354(6352):392–394

    Article  CAS  PubMed  Google Scholar 

  34. Roche PA, Teletski CL, Stang E, Bakke O, Long EO (1993) Cell surface HLA-DR-invariant chain complexes are targeted to endosomes by rapid internalization. Proc Natl Acad Sci USA 90(18):8581–8585

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Wraight CJ, van Endert P, Moller P, Lipp J, Ling NR, MacLennan IC, Koch N, Moldenhauer G (1990) Human major histocompatibility complex class II invariant chain is expressed on the cell surface. J Biol Chem 265(10):5787–5792

    CAS  PubMed  Google Scholar 

  36. Avva RR, Cresswell P (1994) In vivo and in vitro formation and dissociation of HLA-DR complexes with invariant chain-derived peptides. Immunity 1(9):763–774

    Article  CAS  PubMed  Google Scholar 

  37. Denzin LK, Robbins NF, Carboy-Newcomb C, Cresswell P (1994) Assembly and intracellular transport of HLA-DM and correction of the class II antigen-processing defect in T2 cells. Immunity 1(7):595–606

    Article  CAS  PubMed  Google Scholar 

  38. Doebele RC, Busch R, Scott HM, Pashine A, Mellins ED (2000) Determination of the HLA-DM interaction site on HLA-DR molecules. Immunity 13(4):517–527

    Article  CAS  PubMed  Google Scholar 

  39. Stang E, Guerra CB, Amaya M, Paterson Y, Bakke O, Mellins ED (1998) DR/CLIP (class II-associated invariant chain peptides) and DR/peptide complexes colocalize in prelysosomes in human B lymphoblastoid cells. J Immunol 160(10):4696–4707

    CAS  PubMed  Google Scholar 

  40. Guy K, Van Heyningen V, Cohen BB, Deane DL, Steel CM (1982) Differential expression and serologically distinct subpopulations of human Ia antigens detected with monoclonal antibodies to Ia alpha and beta chains. Eur J Immunol 12(11):942–948

    Article  CAS  PubMed  Google Scholar 

  41. Denzin LK, Hammond C, Cresswell P (1996) HLA-DM interactions with intermediates in HLA-DR maturation and a role for HLA-DM in stabilizing empty HLA-DR molecules. J Exp Med 184(6):2153–2165

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Lee AW, Wang N, Hornell TM, Harding JJ, Deshpande C, Hertel L, Lacaille V, Pashine A, Macaubas C, Mocarski ES, Mellins ED (2011) Human cytomegalovirus decreases constitutive transcription of MHC class II genes in mature Langerhans cells by reducing CIITA transcript levels. Mol Immunol 48(9–10):1160–1167

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Hitzel C, Gruneberg U, van Ham M, Trowsdale J, Koch N (1999) Sodium dodecyl sulfate-resistant HLA-DR “superdimer” bands are in some cases class II heterodimers bound to antibody. J Immunol 162(8):4671–4676

    CAS  PubMed  Google Scholar 

  44. Lampson LA, Levy R (1980) Two populations of Ia-like molecules on a human B cell line. J Immunol 125(1):293–299

    CAS  PubMed  Google Scholar 

  45. Guerra CB, Busch R, Doebele RC, Liu W, Sawada T, Kwok WW, Chang MD, Mellins ED (1998) Novel glycosylation of HLA-DRalpha disrupts antigen presentation without altering endosomal localization. J Immunol 160(9):4289–4297

    CAS  PubMed  Google Scholar 

  46. Watanabe M, Suzuki T, Taniguchi M, Shinohara N (1983) Monoclonal anti-Ia murine alloantibodies crossreactive with the Ia-homologues of other mammalian species including humans. Transplantation 36(6):712–718

    Article  CAS  PubMed  Google Scholar 

  47. Carven GJ, Chitta S, Hilgert I, Rushe MM, Baggio RF, Palmer M, Arenas JE, Strominger JL, Horejsi V, Santambrogio L, Stern LJ (2004) Monoclonal antibodies specific for the empty conformation of HLA-DR1 reveal aspects of the conformational change associated with peptide binding. J Biol Chem 279(16):16561–16570

    Article  CAS  PubMed  Google Scholar 

  48. Potolicchio I, Chitta S, Xu X, Fonseca D, Crisi G, Horejsi V, Strominger JL, Stern LJ, Raposo G, Santambrogio L (2005) Conformational variation of surface class II MHC proteins during myeloid dendritic cell differentiation accompanies structural changes in lysosomal MIIC. J Immunol 175(8):4935–4947

    Article  CAS  PubMed  Google Scholar 

  49. Johnson JP, Meo T, Riethmuller G, Schendel DJ, Wank R (1982) Direct demonstration of an HLA-DR allotypic determinant on the low molecular weight (beta) subunit using a mouse monoclonal antibody specific for DR3. J Exp Med 156(1):104–111

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Johnson JP, Wank R (1984) Identification of two cis-encoded HLA-DQ molecules that carry distinct alloantigenic specificities. J Exp Med 160(5):1350–1359

    Article  CAS  PubMed  Google Scholar 

  51. Mellins E, Kempin S, Smith L, Monji T, Pious D (1991) A gene required for class II-restricted antigen presentation maps to the major histocompatibility complex. J Exp Med 174(6):1607–1615

    Article  CAS  PubMed  Google Scholar 

  52. Sanderson F, Thomas C, Neefjes J, Trowsdale J (1996) Association between HLA-DM and HLA-DR in vivo. Immunity 4(1):87–96

    Article  CAS  PubMed  Google Scholar 

  53. Verreck FA, Fargeas CA, Hämmerling GJ (2001) Conformational alterations during biosynthesis of HLA-DR3 molecules controlled by invariant chain and HLA-DM. Eur J Immunol 31(4):1029–1036

    Article  CAS  PubMed  Google Scholar 

  54. Patil NS, Pashine A, Belmares MP, Liu W, Kaneshiro B, Rabinowitz J, McConnell H, Mellins ED (2001) Rheumatoid arthritis (RA)-associated HLA-DR alleles form less stable complexes with class II-associated invariant chain peptide than non-RA-associated HLA-DR alleles. J Immunol 167(12):7157–7168

    Article  CAS  PubMed  Google Scholar 

  55. Hou T, Macmillan H, Chen Z, Keech CL, Jin X, Sidney J, Strohman M, Yoon T, Mellins ED (2011) An insertion mutant in DQA1*0501 restores susceptibility to HLA-DM: implications for disease associations. J Immunol 187(5):2442–2452

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Spits H, Borst J, Giphart M, Coligan J, Terhorst C, De Vries JE (1984) HLA-DC antigens can serve as recognition elements for human cytotoxic T lymphocytes. Eur J Immunol 14(4):299–304

    Article  CAS  PubMed  Google Scholar 

  57. Fallang LE, Roh S, Holm A, Bergseng E, Yoon T, Fleckenstein B, Bandyopadhyay A, Mellins ED, Sollid LM (2008) Complexes of two cohorts of CLIP peptides and HLA-DQ2 of the autoimmune DR3-DQ2 haplotype are poor substrates for HLA-DM. J Immunol 181(8):5451–5461

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Huan J, Meza-Romero R, Mooney JL, Vandenbark AA, Offner H, Burrows GG (2011) Single-chain recombinant HLA-DQ2.5/peptide molecules block alpha2-gliadin-specific pathogenic CD4+ T-cell proliferation and attenuate production of inflammatory cytokines: a potential therapy for celiac disease. Mucosal Immunol 4(1):112–120

    Article  CAS  PubMed  Google Scholar 

  59. Viken HD, Paulsen G, Sollid LM, Lundin KE, Tjonnfjord GE, Thorsby E, Gaudernack G (1995) Characterization of an HLA-DQ2-specific monoclonal antibody. Influence of amino acid substitutions in DQ beta 1*0202. Hum Immunol 42(4):319–327

    Article  CAS  PubMed  Google Scholar 

  60. Shookster L, Matsuyama T, Burmester G, Winchester R (1987) Monoclonal antibody 1a3 recognizes a monomorphic epitope unique to DQ molecules. Hum Immunol 20(1):59–70

    Article  CAS  PubMed  Google Scholar 

  61. Robbins PA, Evans EL, Ding AH, Warner NL, Brodsky FM (1987) Monoclonal antibodies that distinguish between class II antigens (HLA-DP, DQ, and DR) in 14 haplotypes. Hum Immunol 18(4):301–313

    Article  CAS  PubMed  Google Scholar 

  62. Radka SF, Machamer CE, Cresswell P (1984) Analysis of monoclonal antibodies reactive with human class II beta chains by two-dimensional electrophoresis and Western blotting. Hum Immunol 10(3):177–186

    Article  CAS  PubMed  Google Scholar 

  63. Koch N, Koch S, Hämmerling GJ (1982) Ia invariant chain detected on lymphocyte surfaces by monoclonal antibody. Nature 299(5884):644–645

    Article  CAS  PubMed  Google Scholar 

  64. Anderson MS, Miller J (1992) Invariant chain can function as a chaperone protein for class II major histocompatibility complex molecules. Proc Natl Acad Sci USA 89(6):2282–2286

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  65. Germain RN, Hendrix LR (1991) MHC class II structure, occupancy and surface expression determined by post-endoplasmic reticulum antigen binding. Nature 353(6340):134–139. doi:10.1038/353134a0

    Article  CAS  PubMed  Google Scholar 

  66. Mehringer JH, Harris MR, Kindle CS, McCourt DW, Cullen SE (1991) Characterization of fragments of the murine Ia-associated invariant chain. J Immunol 146(3):920–927

    CAS  PubMed  Google Scholar 

  67. Bhattacharya A, Dorf ME, Springer TA (1981) A shared alloantigenic determinant on Ia antigens encoded by the I-A and I-E subregions: evidence for I region gene duplication. J Immunol 127(6):2488–2495

    CAS  PubMed  Google Scholar 

  68. Janeway CA Jr, Conrad PJ, Lerner EA, Babich J, Wettstein P, Murphy DB (1984) Monoclonal antibodies specific for Ia glycoproteins raised by immunization with activated T cells: possible role of T cellbound Ia antigens as targets of immunoregulatory T cells. J Immunol 132(2):662–667

    PubMed  Google Scholar 

  69. Bikoff EK, Germain RN, Robertson EJ (1995) Allelic differences affecting invariant chain dependency of MHC class II subunit assembly. Immunity 2(3):301–310

    Article  CAS  PubMed  Google Scholar 

  70. Oi VT, Jones PP, Goding JW, Herzenberg LA (1978) Properties of monoclonal antibodies to mouse Ig allotypes, H-2, and Ia antigens. Curr Top Microbiol Immunol 81:115–120

    CAS  PubMed  Google Scholar 

  71. Koch N, Hämmerling GJ, Tada N, Kimura S, Hämmerling U (1982) Cross-blocking studies with monoclonal antibodies against I-A molecules of haplotypes b, d and k. Eur J Immunol 12(11):909–914

    Article  CAS  PubMed  Google Scholar 

  72. Dang LH, Lien LL, Benacerraf B, Rock KL (1993) A mutant antigen-presenting cell defective in antigen presentation expresses class II MHC molecules with an altered conformation. J Immunol 150(10):4206–4217

    CAS  PubMed  Google Scholar 

  73. Kappler JW, Skidmore B, White J, Marrack P (1981) Antigen-inducible, H-2-restricted, interleukin-2-producing T cell hybridomas. Lack of independent antigen and H-2 recognition. J Exp Med 153(5):1198–1214

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  74. Steinman RM, Nogueira N, Witmer MD, Tydings JD, Mellman IS (1980) Lymphokine enhances the expression and synthesis of Ia antigens on cultured mouse peritoneal macrophages. J Exp Med 152(5):1248–1261

    Article  CAS  PubMed  Google Scholar 

  75. Ozato K, Mayer NM, Sachs DH (1982) Monoclonal antibodies to mouse major histocompatibility complex antigens. Transplantation 34(3):113–120

    Article  CAS  PubMed  Google Scholar 

  76. Rinderknecht CH, Belmares MP, Catanzarite TL, Bankovich AJ, Holmes TH, Garcia KC, Nanda NK, Busch R, Kovats S, Mellins ED (2007) Posttranslational regulation of I-Ed by affinity for CLIP. J Immunol 179(9):5907–5915

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This work was supported by NIH grants F32 AI089090 to TH and AI095813 and AI28809 to E.D.M. R.B. was supported by a Senior Research Fellowship from Arthritis Research UK (ref. 18543).

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Authors and Affiliations

  1. Department of Pediatrics, Division of Transplantation Biology and Immunology, Stanford University Medical School, Stanford, CA, USA

    Tieying Hou

  2. Division of Transplantation Biology and Immunology, Department of Pediatrics, Stanford University Medical School, Stanford, CA, USA

    Cornelia H. Rinderknecht, Andreas V. Hadjinicolaou & Elizabeth Mellins

  3. Division of Rheumatology, Department of Medicine, University of Cambridge, Addenbrooke’s Hospital, Cambridge, UK

    Robert Busch

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  1. Tieying Hou
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  2. Cornelia H. Rinderknecht
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  3. Andreas V. Hadjinicolaou
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  4. Robert Busch
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  5. Elizabeth Mellins
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Correspondence to Elizabeth Mellins .

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  1. Institut National de la Santé et de la Recherche Médicale, Unité 1013, Université Paris Descartes, Faculté de médecine, Paris, France

    Peter van Endert

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Hou, T., Rinderknecht, C.H., Hadjinicolaou, A.V., Busch, R., Mellins, E. (2013). Pulse–Chase Analysis for Studies of MHC Class II Biosynthesis, Maturation, and Peptide Loading. In: van Endert, P. (eds) Antigen Processing. Methods in Molecular Biology™, vol 960. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-218-6_31

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  • DOI: https://doi.org/10.1007/978-1-62703-218-6_31

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