Designing, Constructing and Immunogenic Evaluation of
Polytope DNA Constructs by the Application of Hepatitis
C Virus Immunodominant Epitopes in BALB/c Mice
Arash Memarnejadian, Ph.D., Farzin Roohvand, Ph.D.*, Arash Arashkia, Ph.D., Farideh Berjisian, B.Sc., Mohammad Reza Aghasadeghi, Ph.D.
Hepatitis and AIDS Department, NRGB Lab, Pasteur Institute of Iran, Tehran, Iran
* Corresponding Address: P.O.Box: 1316943551, Hepatitis and AIDS Department, NRGB Lab, Pasteur Institute of Iran, Tehran, Iran
Email: rfarzin@pasteur.ac.ir
Received: 26/May/2008, Accepted: 11/Nov/2008 Abstract
Objective: Polytope DNA vaccines, capable of focusing the cytotoxic T lymphocyte (CTL)
response on critical epitopes, represent a promising approach in HCV immunotherapy. Nev-ertheless, due to controversial rules governing epitope processing and the low level expres-sion/immunogenicity of recombinant polytope peptides, designing and primary expression/ immunogenicity analysis of these vaccine types should be the first consideration prior to costly transgenic animal studies.
Materials and Methods: Four HLA-A2 and H-2d restricted CTL epitopes were selected and
designed in three appropriate sequential tandems based on epitope and proteasomal cleav-age predictions. The related nucleotide sequences were synthesized using SOEing PCR method and cloned into a pcDNA3.1 vector, either alone or fused to the small hepatitis B sur-face antigen (HBsAg-S) gene. Following the preparation of polyclonal anti-sera, expression/ secretion of polytopes was evaluated in Cos-7 cells by using immunofluorescence, Western-blot, dot Western-blot, ELISA and RT-PCR techniques. The immunogenicity of the plasmids was also assessed through the delayed-type hypersensitivity (DTH) assay in BALB/c mice.
Results: Due to in silico designs and optimizations, the polytope products of constructed
plasmids were efficiently detected in vitro through common techniques and HBsAg-S-based
particles were shown to be secreted into the culture media (up to 30%). Moreover, all plas-mids were able to efficiently induce a positive DTH response while HBsAg-S fusion constructs indicated a significant immunopotential effect towards the incorporated mouse epitopes.
Conclusion: Designed polytope constructs of this study are efficiently expressed and
proc-essed. They have the required initial potency for further immunogenicity analysis in trans-genic mice.
Keywords: Epitope, Hepatitis C Virus, DNA Vaccine, Cytotoxic T-Lymphocytes
ﺯﺍ
ﻩﺩﺎﻔﺘﺳﺍ
ﺎﺑ
پﻮﺗ
ﻰﻠﭘ
DNA
ﻯﺎﻫ
ﻩﺯﺎﺳ
ﻰﻳﺍﺯ
ﻰﻨﻤﻳﺍ
ﻰﺑﺎﻳﺯﺭﺍ
ﻭ
ﺖﺧﺎﺳ
،ﻰﺣﺍﺮﻃ
BALB/c
ﺵﻮﻣ
ﺭﺩ
ﻰﺳ
ﺖﻴﺗﺎﭙﻫ
ﺱﻭﺮﻳﻭ
ﺐﻟﺎﻏ
ﻯﺎﻫ
پﻮﺗ
ﻰﭘﺍ
،،Ph.D.ﻰﻗﺩﺎﺻﺎﻗﺁﺎﺿﺭﺪﻤﺤﻣ،،،B.Sc.ﻥﺎﻴﺴﻴﺟﺮﺑﻩﺪﻳﺮﻓ،،،Ph.D.ﺎﻴﻛﺵﺭﺁﺵﺭﺁ، *
،Ph.D.ﺪﻧﻮﺣﻭﺭﻦﻳﺯﺮﻓ،،Ph.D.ﻥﺎﻳﺩﺍﮋﻧﺭﺎﻤﻌﻣﺵﺭﺁ
ﻥﺍﺮﻳﺍ،ﻥﺍﺮﻬﺗ،ﺐﻴﻛﺮﺗﻮﻧﻥژﻚﻧﺎﺑ،ﺯﺪﻳﺍﻭﺖﻴﺗﺎﭙﻫﺶﺨﺑ،ﻥﺍﺮﻳﺍﺭﻮﺘﺳﺎﭘﻮﺘﻴﺘﺴﻧﺍ
ﺐﻴﻛﺮﺗﻮﻧﻥژﻚﻧﺎﺑ،ﺯﺪﻳﺍﻭﺖﻴﺗﺎﭙﻫﺶﺨﺑ،ﻥﺍﺮﻳﺍﺭﻮﺘﺳﺎﭘﻮﺘﻴﺘﺴﻧﺍ ،1316943551 :ﻰﺘﺴﭘﻕﻭﺪﻨﺻ،ﻥﺍﺮﻬﺗ،ﻥﺍﺮﻳﺍ :ﻝﻮﺌﺴﻣﻩﺪﻨﺴﻳﻮﻧﺱﺭﺩﺁ *
Email: rfarzin@pasteur.ac.ir:ﻚﻴﻧﻭﺮﺘﻜﻟﺍﺖﺴﭘ
87/8/21 :ﻪﻟﺎﻘﻣﺵﺮﻳﺬﭘ،87/3/6 :ﻪﻟﺎﻘﻣﺖﻓﺎﻳﺭﺩ
ﻩﺪﻴﻜﭼ
ﺖﻴﺗﺎﭙﻫﺱﻭﺮﻳﻭﺐﻟﺎﻏﻱﺎﻫپﻮﺗﻲﭘﺍﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑپﻮﺗﻰﻠﭘ DNA ﻱﺎﻫﻩﺯﺎﺳﻲﻳﺍﺯﻲﻨﻤﻳﺍﻲﺑﺎﻳﺯﺭﺍﻭﺖﺧﺎﺳ،ﻲﺣﺍﺮﻃ :ﻑﺪﻫ *
BALB/cﺵﻮﻣﺭﺩﻲﺳ
ﻰﻳﺎﺳﺎﻨﺷﻯﺎﻫﻢﺘﻳﺭﻮﮕﻟﺍﺱﺎﺳﺍﺮﺑﻭﺏﺎﺨﺘﻧﺍ H-2dﻭ HLA-A2 ﻪﺑﻪﺘﺴﺑﺍﻭ CTL پﻮﺗﻰﭘﺍﺭﺎﻬﭼ
:ﺎﻫﺵﻭﺭﻭﺩﺍﻮﻣ *
-SOE ﺵﻭﺭ ﺎﺑﻪﻃﻮﺑﺮﻣ ﻯﺪﻴﺗﻮﺌﻠﻛﻮﻧﻯﺎﻫﻰﻟﺍﻮﺗ .ﺪﻧﺪﺷ ﻰﺣﺍﺮﻃ ﻒﻠﺘﺨﻣﺐﻴﺗﺮﺗ ﻪﺳ ﺭﺩﻡﻭﺯﺎﺌﺗﻭﺮﭘﺵﺮﺑ ﻭ پﻮﺗﻰﭘﺍ (HBsAg-S)ﻰﺑﺖﻴﺗﺎﭙﻫﺱﻭﺮﻳﻭﻰﺤﻄﺳﻥژﻰﺘﻧﺁﻥژ ﻪﺑﻞﺼﺘﻣﺮﻴﻏﺎﻳﻞﺼﺘﻣﺕﺭﻮﺻﻪﺑﻭﻩﺪﺷﻪﺘﺧﺎﺳ ing PCR
ﻯﺎﻫﻝﻮﻠﺳ ﺭﺩﺎﻫپﻮﺗﻰﻠﭘ ﺢﺷﺮﺗﻭﻥﺎﻴﺑﻝﺎﻧﻮﻠﻛﻰﻠﭘ ﻡﺮﺳﻰﺘﻧﺁ ﻪﻴﻬﺗﻝﺎﺒﻧﺩ ﻪﺑ .ﺪﻧﺪﺷ ﻥﻮﻠﻛ pcDNA3.1 ﺪﻴﻤﺳﻼﭘﺭﺩ
ﻰﻳﺍﺯﻰﻨﻤﻳﺍ ﻭ ﻰﺳﺭﺮﺑ RT-PCR ﻭ ﺍﺰﻳﻻﺍ ،ﺕﻼﺑ ﺕﺍﺩ ،ﺕﻼﺑ ﻥﺮﺘﺳﻭ ،ﺲﻧﺎﺳﺭﻮﻠﻓﻮﻧﻮﻤﻳﺍ ﻯﺎﻫﺵﻭﺭ ﺎﺑ Cos-7 BALB/cﺵﻮﻣﺭﺩ (Delayed-type Hypersensitivity; DTH) ﻯﺮﻴﺧﺎﺗﺖﻴﺳﺎﺴﺣﺩﺎﻳﺩﺯﺍﻥﻮﻣﺯﺁﺎﺑﺎﻫﻩﺯﺎﺳ .ﺪﺷﻰﺑﺎﻳﺯﺭﺍ
ﺭﺎﻛﻪﺑﺕﺍﺪﻴﻬﻤﺗﻭin silicoﻰﺣﺍﺮﻃﻪﻄﺳﺍﻭﻪﺑﻩﺪﺷﻪﺘﺧﺎﺳﻯﺎﻫﺪﻴﻤﺳﻼﭘﻥﺎﻴﺑﺯﺍﻞﺻﺎﺣپﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﺘﭙﭘ :ﺎﻫﻪﺘﻓﺎﻳ *
ﻪﺑ HBsAg-Sﺎﺑﺪﻳﺮﺒﻴﻫﺕﺍﺭﺫﺢﺷﺮﺗﻭﻩﺪﺷﻰﻳﺎﺳﺎﻨﺷﻞﻣﺎﻛﺭﻮﻃﻪﺑﻩﺎﮕﺸﻳﺎﻣﺯﺁﻂﻴﺤﻣﺭﺩﻝﻮﻤﻌﻣﻯﺎﻫﻚﻴﻨﻜﺗﺎﺑ،ﻪﺘﻓﺭ
ﺪﻧﺩﻮﺑ DTH ﺦﺳﺎﭘﺪﻣﺁﺭﺎﻛﻚﻳﺮﺤﺗﻪﺑﺭﺩﺎﻗﺎﻫﺪﻴﻤﺳﻼﭘﻪﻴﻠﻛﻦﻴﻨﭼﻢﻫ .ﺪﺷﻩﺩﺍﺩﻥﺎﺸﻧ (ﺪﺻﺭﺩ 30 ﻥﺍﺰﻴﻣﻪﺑ) ﺖﺸﻛﻂﻴﺤﻣ
ﻰﺷﻮﻣﻯﺎﻫپﻮﺗﻰﭘﺍﻪﻴﻠﻋﻩﺪﺷﺩﺎﺠﻳﺍﺦﺳﺎﭘﺮﺑﺍﺭﻯﺭﺍﺩﻰﻨﻌﻣﻩﺪﻨﻳﺍﺰﻓﺮﺛﺍ HBsAg-Sﺎﺑﺪﻳﺮﺒﻴﻫﻯﺎﻫﻩﺯﺎﺳﻥﺎﻴﻣﻦﻳﺍﺭﺩﻪﻛ .ﺪﻧﺩﺍﺩﻥﺎﺸﻧ
ﺮﺘﺸﻴﺑﻰﺳﺭﺮﺑﺭﻮﻈﻨﻣﻪﺑﻡﺯﻻﻪﻴﻟﻭﺍﻰﻳﺍﺭﺎﻛﺯﺍﻭﻩﺪﺷﺵﺯﺍﺩﺮﭘﻭﻥﺎﻴﺑﻰﺑﻮﺧﻪﺑﻩﺪﺷﻪﺘﺧﺎﺳپﻮﺗﻰﻠﭘﻯﺎﻫﻩﺯﺎﺳ :ﻱﺮﻴﮔﻪﺠﻴﺘﻧ * .ﺪﺷﺎﺑﻰﻣﺭﺍﺩﺭﻮﺧﺮﺑﻪﺘﺨﻳﺭﺍﺮﺗﺵﻮﻣﺭﺩﻰﺋﺍﺯﻰﻨﻤﻳﺍ
ﻚﻴﺴﻛﻮﺗﻮﺘﻴﺳ T ﻯﺎﻫﺖﻴﺳﻮﻔﻨﻟ،DNA ﻦﺴﻛﺍﻭ،ﻰﺳﺖﻴﺗﺎﭙﻫﺱﻭﺮﻳﻭ،پﻮﺗﻰﭘﺍ:ﻥﺎﮔژﺍﻭﺪﻴﻠﻛ *
122-133 :ﺕﺎﺤﻔﺻ،88 ﻥﺎﺘﺴﺑﺎﺗ،2 ﻩﺭﺎﻤﺷ،ﻢﻫﺩﺯﺎﻳﻝﺎﺳ،ﻪﺘﺧﺎﻳﻰﻜﺷﺰﭘﻪﻣﺎﻨﻠﺼﻓ
ﻪﻣﺪﻘﻣ
Cytotoxic) ﻚﻴﺴﻛﻮﺗﻮﺘﻳﺎﺳ T ﻯﺎﻫﺖﻴﺳﻮﻔﻨﻟ ﻯﺪﻴﻠﻛ ﺶﻘﻧ
ﻰﺳﻭﺮﻳﻭ ﻯﺎﻫﺖﻧﻮﻔﻋ ﻪﻴﻠﻋ ﻉﺎﻓﺩ ﺭﺩ (T-Lymphocytes; CTL
ﻰﺳ ﺖﻴﺗﺎﭙﻫ ﺱﻭﺮﻳﻭ ﺪﻨﻧﺎﻣ ،ﻚﻴﺗﺎﭘﻮﺘﻳﺎﺳﺮﻴﻏ ﻯﺎﻫﺱﻭﺮﻳﻭ ﻩﮋﻳﻭ ﻪﺑ ﺖﺳﺍﻩﺪﺷﻩﺩﺍﺩﻥﺎﺸﻧﻒﻠﺘﺨﻣﺕﺎﻌﻟﺎﻄﻣﺭﺩ (Hepatitis C Virus; ﻭHCV)
ﺖﻴﻤﻫﺍﺯﺍﻰﻛﺎﺣﺰﻴﻧHCVﺖﻧﻮﻔﻋﻩﺪﺷﻥﺎﻣﺭﺩﺩﺭﺍﻮﻣﻰﺳﺭﺮﺑﺍﺮﻴﺧﺍ .(1-4) ﻢﻬﻣﻯﺎﻫپﻮﺗﻰﭘﺍﻪﻴﻠﻋT CD8+ﻯﺎﻫﺖﻴﺳﻮﻔﻨﻟﻪﺒﻧﺎﺟﺪﻨﭼﻭﻯﻮﻗﺦﺳﺎﭘ ﻭﺪﺑﺭﺩﺖﻧﻮﻔﻋﻯﺯﺎﺳﻙﺎﭘﺭﺩﺎﻫﻝﻮﻠﺳﻦﻳﺍﻰﻳﺎﻧﺍﻮﺗﻭﺱﻭﺮﻳﻭﺮﻴﻐﺘﻣﺎﻧﻭ ﺱﻭﺮﻳﻭﻒﻠﺘﺨﻣﻯﺎﻫﻪﻳﻮﺳﺩﻮﺟﻭﻞﻴﻟﺩﻪﺑﻝﺎﺣﻦﻳﺍﺎﺑ .(7-5) ﺖﺳﺍﻉﻭﺮﺷ ﺮﺼﺤﻨﻣﻰﻳﺎﻧﺍﻮﺗﻦﻴﻨﭼﻢﻫﻭ (9 ،8) ﺎﻬﻧﺁﻚﻴﻧژﻰﺘﻧﺁﺭﺎﺘﺧﺎﺳﻉﻮﻨﺗﻭﻭHCV
ﺭﺍﺮﻓﻪﺑﺮﺠﻨﻣﻪﻛ (ﻥﻮﻴﺳﺎﺗﻮﻣ) ﻰﭘﻮﺗﻰﭘﺍﺭﺎﺘﺧﺎﺳﻊﻳﺮﺳﺮﻴﻴﻐﺗﺭﺩﺱﻭﺮﻳﻭﺩﺮﻓﻪﺑ ﻰﻧﺎﻣﺭﺩﻯﺎﻫﻦﺴﻛﺍﻭﺩﺮﺑﺭﺎﻛ،(10 ،3) ﺩﺩﺮﮔﻰﻣﻰﻨﻤﻳﺍﻢﺘﺴﻴﺳﺖﺳﺩﺯﺍﻥﺁ ﻩﺪﺸﻧﻊﻗﺍﻭﺮﺛﻮﻣﻥﻮﻨﻛﺎﺗT CD8+ﻯﺎﻫﻝﻮﻠﺳﺭﺩﻰﺨﺳﺎﭘﻦﻴﻨﭼﺩﺎﺠﻳﺍﻯﺍﺮﺑ ﻪﻛ(DNA Vaccine)ﻚﻴﺳﻼﻛDNAﻯﺎﻫﻦﺴﻛﺍﻭﺮﺿﺎﺣﻝﺎﺣﺭﺩ .ﺖﺳﺍ ﻩﺩﺎﻔﺘﺳﺍﻰﻨﻤﻳﺍﻢﺘﺴﻴﺳﻚﻳﺮﺤﺗﺖﻬﺟﻢﺴﻴﻧﺎﮔﺭﺍﻭﺮﻜﻴﻣﻞﻣﺎﻛﻥژﻰﺘﻧﺁﺯﺍ ﻯﺎﻫﻝﻮﻠﺳﻚﻳﺮﺤﺗﻭTh1ﺦﺳﺎﭘﺎﻘﻟﺍﻯﺎﻫﻩﺍﺭﺯﺍﻰﻜﻳﻥﺍﻮﻨﻋﻪﺑ،ﺪﻨﻨﻛﻰﻣ
ﺩﺭﻮﻣﺭﺩﻪﭼﺮﮔﺍﻦﺴﻛﺍﻭﻉﻮﻧﻦﻳﺍ .(12 ،11) ﺪﻧﺍﻩﺪﺷﻪﺘﻓﺮﻳﺬﭘ CTLs
ﺭﺩ ﺍﺭﻯﺮﻴﮔﻢﺸﭼ ﻰﻳﺍﺭﺎﻛ ﺎﻣﺍ ،ﺖﺳﺍ ﻩﺩﻮﺑ ﻖﻓﻮﻣ ﺎﻫﻥژﻮﺗﺎﭘﺯﺍ ﻰﺧﺮﺑ ﻩﺩﺍﺪﻧﻥﺎﺸﻧﺩﻮﺧﺯﺍHCVﺪﻨﻧﺎﻣﻦﻣﺰﻣﻰﺳﻭﺮﻳﻭﻯﺎﻫﺖﻧﻮﻔﻋﻯﺯﺎﺳﻙﺎﭘ
ﺩﺍﺪﻌﺗ ﻥﺩﻮﺑ ﻰﻓﺎﻛﺎﻧ ﻪﺑ ﻥﺍﻮﺗﻰﻣ ﻯﺪﻣﺁﺭﺎﻛﺎﻧ ﻦﻳﺍﻞﻠﻋ ﻪﻠﻤﺟ ﺯﺍ .ﺖﺳﺍ
ﺎﻳ (13) ﻥژﻰﺘﻧﺁ ﻚﻳﺭﺩ (Protective) ﻆﻓﺎﺤﻣ CTLﻯﺎﻫپﻮﺗﻰﭘﺍ
ﺭﺩﻩﻭﻼﻋﻪﺑ .ﺩﺮﻛﻩﺭﺎﺷﺍ (14-16) ﺱﻭﺮﻳﻭﺭﺍﺮﻓﻪﺠﻴﺘﻧﺭﺩﻭﻥﻮﻴﺳﺎﺗﻮﻣ ﻥژﻰﺘﻧﺁﺭﺩﺩﻮﺟﻮﻣﻩﺪﻨﻨﻛﺏﻮﻛﺮﺳﻭﻂﺒﺗﺮﻣﺮﻴﻏﻯﺎﻫپﻮﺗﻰﭘﺍﻯﺩﺭﺍﻮﻣ ﺩﺎﺠﻳﺍﺯﺍﻊﻧﺎﻣﺪﻨﻧﺍﻮﺗﻰﻣﻩﺪﻨﻨﻛﺖﻈﻓﺎﺤﻣﻯﺎﻫپﻮﺗﻰﭘﺍﺎﺑﻞﺧﺍﺪﺗﺖﻠﻋﻪﺑ ﺭﺩﻪﺘﺳﺍﻮﺧﺎﻧﻯﺎﻫﺦﺳﺎﭘﻦﺘﺨﻴﮕﻧﺍﺮﺑﺐﺟﻮﻣﻰﺘﺣﻭﻪﺘﺸﮔﺐﺳﺎﻨﻣﺦﺳﺎﭘ
HCVﺱﻭﺮﻳﻭﺩﺭﻮﻣﺭﺩﺺﺨﺸﻣﺭﻮﻃﻪﺑ .(17) ﺪﻧﺩﺮﮔﻥﻮﻴﺳﺎﻨﻴﺴﻛﺍﻭﺮﺛﺍ ﻯﺎﻫﻥژﻰﺘﻧﺁ ﺯﺍ ﻰﺧﺮﺑ ﺩﺭﻮﻣ ﺭﺩ ﻰﻨﻤﻳﺍﻢﺘﺴﻴﺳ ﻰﮔﺪﻨﻨﻛﺏﻮﻛﺮﺳ ﺮﺛﺍ ﻥﺍﻮﻨﻋﻪﺑﻪﻛﺖﺳﺍﻩﺪﺷﻩﺪﻫﺎﺸﻣNS3ﻭCore ،E1 ،E2ﺪﻨﻧﺎﻣﺱﻭﺮﻳﻭ
، Tﻯﺎﻫﻝﻮﻠﺳﺪﻨﻧﺎﻣﺎﻫﻝﻮﻠﺳﺯﺍﻰﺧﺮﺑﺩﺮﻜﻠﻤﻋﻒﻴﻌﻀﺗﻯﺍﺮﺑﻰﻤﺴﻴﻧﺎﻜﻣ
ﺭﺎﺘﺧﺎﺳﺯﺍﻩﺩﺎﻔﺘﺳﺍﺮﻣﺍﻦﻴﻤﻫ .(18-21) ﺩﻮﺷﻰﻣﺮﻛﺫﻚﻴﺘﻳﺭﺪﻧﺩﻭNK
.ﺩﺯﺎﺳﻰﻣﺩﻭﺪﺤﻣﻥﻮﻴﺳﺎﻨﻴﺴﻛﺍﻭﺭﻮﻈﻨﻣﻪﺑﺍﺭﺎﻫﻥژﻰﺘﻧﺁﻦﻳﺍﻰﻌﻴﺒﻃ
Polytope DNA)پﻮﺗﻰﻠﭘﺎﻳﻰﭘﻮﺗﻰﭘﺍﺪﻨﭼDNA ﻯﺎﻫﻦﺴﻛﺍﻭ
ﻢﻫﺍﺮﻓ ﺍﺭ ﻥﺎﻜﻣﺍ ﻦﻳﺍﻦﻳﺰﮕﻳﺎﺟ ﻯﮋﺗﺍﺮﺘﺳﺍ ﻚﻳﻥﺍﻮﻨﻋ ﻪﺑ (Vaccines
ﺭﻮﻛﺬﻣﻯﺎﻫﻦﺴﻛﺍﻭﺐﻴﺗﺮﺗﻦﻳﺍﻪﺑ .(23 ،22) ﺩﺩﺮﮔﻯﺮﻴﮔﻮﻠﺟﺰﻴﻧﺮﻀﻣ ﻯﺎﻫﺖﻧﻮﻔﻋﻰﺧﺮﺑﺩﺭﻮﻣﺭﺩﺎﻬﻧﺁﻪﺑﺖﺒﺴﻧﻩﺪﺷﺩﺎﺠﻳﺍﻰﻨﻤﻳﺍﺦﺳﺎﭘﻪﻛ
ﻪﺑ ،(24-26) ﺖﺳﺍ ﻩﺪﺷ ﻩﺩﺍﺩ ﻥﺎﺸﻧ ﻰﺑﻮﺧ ﻪﺑ ﺎﻫﺭﻮﻣﻮﺗ ﻭ ﻰﺳﻭﺮﻳﻭ
ﺡﺮﻄﻣﺮﻄﺧﻰﺑﻭﺪﻣﺁﺭﺎﻛDNAﻯﺎﻫﻦﺴﻛﺍﻭﺯﺍﻯﺪﻳﺪﺟﻞﺴﻧﻥﺍﻮﻨﻋ
ﻯﺎﻫﻞﺣﻩﺍﺭﻦﺘﻓﺎﻳﺪﻨﻣﺯﺎﻴﻧﻦﺴﻛﺍﻭﻉﻮﻧﻦﻳﺍﻪﻌﺳﻮﺗﺩﻮﺟﻭﻦﻳﺍﺎﺑ .ﺪﻨﺷﺎﺑﻰﻣ ﻰﻳﺍﺯﻰﻨﻤﻳﺍﺶﻳﺍﺰﻓﺍ،ﻰﻋﻮﻨﺼﻣﻯﺎﻫﻰﻟﺍﻮﺗﻦﻳﺍﻥﺎﻴﺑﺶﻳﺍﺰﻓﺍﻯﺍﺮﺑﺐﺳﺎﻨﻣ ﻦﺴﻛﺍﻭﻰﻳﺍﺭﺎﻛ ﺶﺠﻨﺳﺖﻬﺟﺐﺳﺎﻨﻣﻯﺎﻫﺵﻭﺭ ﻯﺭﺍﺬﮔﻪﻳﺎﭘﻭﺎﻬﻧﺁ ﻯﺎﻫﺪﻴﺘﭙﭘ ﻪﻛ ﺖﺳﺍ ﺮﻛﺫ ﻪﺑ ﻡﺯﻻ .ﺪﺷﺎﺑﻰﻣ ﻰﻫﺎﮕﺸﻳﺎﻣﺯﺁ ﻥﺍﻮﻴﺣ ﺭﺩ ﻰﻤﻛﻯﺭﺍﺪﻳﺎﭘ ﺎﻣﻮﻤﻋ ﻡﻮﺳﻭﻡﻭﺩ ﻥﺎﻤﺘﺧﺎﺳﻥﺍﺪﻘﻓ ﺖﻠﻋ ﻪﺑ پﻮﺗﻰﻠﭘ ،27 ،23) ﺪﻨﺷﺎﺑﻰﻣﺱﺎﺴﺣﻪﻤﺟﺮﺗﻦﻴﺣﺭﺩﺰﻴﻟﻮﺌﺗﻭﺮﭘﻪﺑﺖﺒﺴﻧﻭﻪﺘﺷﺍﺩ ﻢﻴﻘﺘﺴﻣﺮﻴﻏﺭﻮﻃﻪﺑﺪﻧﺍﻮﺗﻰﻣﺎﻫﺪﻴﺘﭙﭘﻦﻳﺍﺩﻭﺪﺤﻣﻥﺎﻴﺑﻪﻛﺎﺠﻧﺁﺯﺍ .(28 ﻉﻮﻧ ﻦﻳﺍﻥﺎﻴﺑﻰﻫﺎﮕﺸﻳﺎﻣﺯﺁﻰﺑﺎﻳﺯﺭﺍ،ﺩﺭﺍﺬﮕﺑ ﺮﻴﺛﺎﺗﺎﻬﻧﺁ ﻰﻳﺍﺯﻰﻨﻤﻳﺍﺮﺑ ﻯﺮﺘﺸﻴﺑﺖﻴﻤﻫﺍﺯﺍﻚﻴﺳﻼﻛDNAﻯﺎﻫﻦﺴﻛﺍﻭﺎﺑﻪﺴﻳﺎﻘﻣﺭﺩﻦﺴﻛﺍﻭ
ﻰﻀﻴﻘﻧﻭﺪﺿﺕﺎﻋﻼﻃﺍﺮﺿﺎﺣﻝﺎﺣﺭﺩﻦﻴﻨﭼﻢﻫ .ﺪﺷﺎﺑﻰﻣﺭﺍﺩﺭﻮﺧﺮﺑ ﺎﻬﻧﺁﻝﺎﺼﺗﺍﻰﺘﻴﻨﻴﻓﺍ،ﺎﻫپﻮﺗﻰﭘﺍﻑﺩﺍﺮﺗﻭﻯﺮﻴﮔﺭﺍﺮﻗﻞﺤﻣﺮﻴﺛﺎﺗﺩﺭﻮﻣﺭﺩ (Th) ﻰﻜﻤﻛﻯﺎﻫپﻮﺗﻰﭘﺍﺩﻮﺟﻭﻡﺪﻋﺎﻳﺩﻮﺟﻭﻭMHCﻝﻮﻜﻟﻮﻣﻪﺑ
ﻪﻛ (30 ،29 ،27) ﺖﺳﺍﺖﺳﺩﺭﺩپﻮﺗﻰﻠﭘﻯﺎﻫﻦﺴﻛﺍﻭﻰﻳﺍﺯﻰﻨﻤﻳﺍﺮﺑ
ﻪﺑﻩﺭﺎﺷﺍﺖﻳﺎﻬﻧﺭﺩ .ﺪﺷﺎﺑﻰﻣﺮﺘﺸﻴﺑﺕﺎﻘﻴﻘﺤﺗﻡﺎﺠﻧﺍﺪﻨﻣﺯﺎﻴﻧﺎﻬﻧﺁﻞﻴﻤﻜﺗ پﻮﺗﻰﻠﭘﻯﺎﻫﻦﺴﻛﺍﻭﻰﻳﺍﺯﻰﻨﻤﻳﺍﻰﺳﺭﺮﺑﻪﻛﺖﺳﺍﻯﺭﻭﺮﺿﻪﺘﻜﻧﻦﻳﺍ
ﻭ
(HLA) ﻯﺍﺭﺍﺩ ﻪﺘﺨﻳﺭﺍﺮﺗ ﺕﺎﻧﺍﻮﻴﺣﺯﺍ ﻩﺩﺎﻔﺘﺳﺍ ﻪﺑ ﺯﺎﻴﻧﻞﻴﻟﺩ ﻪﺑ ﻰﻧﺎﺴﻧﺍ
ﻦﻳﺍﺮﻴﺜﻜﺗﻭﻪﻴﻬﺗﺕﻼﻜﺸﻣﻭﻰﻧﺎﺴﻧﺍHuman Leukocyte Antigen
ﺪﺷﺎﺑﻰﻤﻧﻚﻴﺳﻼﻛﻯﺎﻫﻦﺴﻛﺍﻭﻰﺘﺣﺍﺭﻪﺑﻭﻩﺩﻮﺑﺮﺑﻪﻨﻳﺰﻫ،ﻥﺍﻮﻴﺣﻉﻮﻧ ﻯﮋﺗﺍﺮﺘﺳﺍﻪﻌﺳﻮﺗﻪﻛﺖﻓﺮﮔﻪﺠﻴﺘﻧﻥﺍﻮﺗﻰﻣﻕﻮﻓﺩﺭﺍﻮﻣﻪﺑﻪﺟﻮﺗﺎﺑ .(31) ﻪﭼﺮﻫﻰﺣﺍﺮﻃﺎﺘﺳﺍﺭﻦﻳﺍﺭﺩﻭﻩﺩﻮﺑﺮﺘﺸﻴﺑﻱﺎﻫﻖﻴﻘﺤﺗﺪﻨﻣﺯﺎﻴﻧپﻮﺗﻰﻠﭘ ﻡﺎﺠﻧﺍﺯﺍﻞﺒﻗﻦﺴﻛﺍﻭﻰﻳﺍﺯﻰﻨﻤﻳﺍﻭﻥﺎﻴﺑﺯﺍﻪﻴﻟﻭﺍﻥﺎﻨﻴﻤﻃﺍﺰﻴﻧﻭﺮﺗﻖﻴﻗﺩ ﺶﻫﺎﻛﻪﺑﺪﻧﺍﻮﺗﻰﻣ،ﻪﺘﺨﻳﺭﺍﺮﺗﻥﺍﻮﻴﺣﻯﻭﺭﺮﺑﻰﻨﻴﻟﺎﺑﺖﺤﺗﺕﺎﺸﻳﺎﻣﺯﺁ
.ﺩﻮﺷﺮﺠﻨﻣﺎﻫﻪﻨﻳﺰﻫﻭﻥﺎﻣﺯ
ﻥﺍﻮﻨﻋﻪﺑ) ﻰﺳﺖﻴﺗﺎﭙﻫﺱﻭﺮﻳﻭNS3ﻭCore ،E2ﻯﺎﻫﻥژﻰﺘﻧﺁ
ﻰﻳﺎﻫﻰﮔﮋﻳﻭﻥﺩﻮﺑﺍﺭﺍﺩﺖﻠﻋﻪﺑ (ﻦﻣﺰﻣﺖﻴﺗﺎﭙﻫﻞﻣﺎﻋﻥژﻮﺗﺎﭘﻦﻳﺮﺗﻢﻬﻣ ﺱﺮﺘﺳﺩ ﺭﺩ ،(Core) ﻰﻳﺍﺯﻯﺭﺎﻤﻴﺑ ﺭﺩ ﺖﻟﺎﺧﺩ ﻭ ﻥﺩﻮﺑ ﺮﻴﻐﺘﻣﺎﻧ ﺮﻴﻈﻧ ﺭﺩ ﺖﻟﺎﺧﺩ ﻦﻴﻨﭼﻢﻫ (E2) ﺪﺒﻛ ﻯﺎﻫﻝﻮﻠﺳ ﻪﺑ ﺱﻭﺮﻳﻭﻝﺎﺼﺗﺍ ﻥﺩﻮﺑ ﻯﺯﺎﺳﻙﺎﭘﺎﺑﻩﺪﺷﺩﺎﺠﻳﺍﻰﻨﻤﻳﺍﺦﺳﺎﭘﻁﺎﺒﺗﺭﺍﻩﺪﻫﺎﺸﻣﻭﺱﻭﺮﻳﻭﺮﻴﺜﻜﺗ ﺭﺩﻩﺩﺎﻔﺘﺳﺍﻯﺍﺮﺑﺐﺳﺎﻨﻣﻯﺎﻫﻥژﻰﺘﻧﺁﻥﺍﻮﻨﻋﻪﺑﻩﺭﺍﻮﻤﻫ،(NS3) ﺖﻧﻮﻔﻋ
.(33 ،32) ﺪﻧﺍﻩﺩﻮﺑﻪﺟﻮﺗﺩﺭﻮﻣﻰﻧﺎﻣﺭﺩﻯﺎﻫﻦﺴﻛﺍﻭ
ﺪﻨﭼﻯﺎﻫﻦﺴﻛﺍﻭﻯﮋﺗﺍﺮﺘﺳﺍﻰﻳﺍﺭﺎﻛﺕﺎﺒﺛﺍﺮﺿﺎﺣﻖﻴﻘﺤﺗﺯﺍﻑﺪﻫ ﺯﺍﻪﺘﻓﺮﮔﺮﺑCTLپﻮﺗﻰﭘﺍﺭﺎﻬﭼﻪﻴﻠﻋﻰﻨﻤﻳﺍﺦﺳﺎﭘﺩﺎﺠﻳﺍﺭﺩﻰﭘﻮﺗﻰﭘﺍ
ﻦﻳﺍﺎﺑ ﻂﺒﺗﺮﻣﺕﻻﻮﻬﺠﻣﻰﺧﺮﺑﻰﺳﺭﺮﺑﻦﻴﻨﭼﻢﻫﻭ ﺭﻮﻛﺬﻣﻪﻴﺣﺎﻧﻪﺳ ﻥژﻰﺘﻧﺁﻩﺪﻨﻳﺍﺰﻓﺍﺮﺛﺍﺰﻴﻧ ﻭﺎﻫپﻮﺗﻰﭘﺍﻑﺩﺍﺮﺗﺮﻴﺛﺎﺗﺪﻨﻧﺎﻣﻦﺴﻛﺍﻭﻉﻮﻧ ﺎﻬﻧﺁﻰﻳﺍﺯﻰﻨﻤﻳﺍﺮﺑ (HBsAg-S) ﻰﺑﺖﻴﺗﺎﭙﻫﺱﻭﺮﻳﻭﻰﺤﻄﺳﻚﭼﻮﻛ ﻚﻴﺗﺎﻣﺭﻮﻔﻧﺍﻮﻧﻮﻤﻳﺍﻯﺎﻫﺰﻴﻟﺎﻧﺁﺯﺍﻯﺮﻴﮔﻩﺮﻬﺑﺎﺑﺭﻮﻈﻨﻣﻦﻳﺍﻯﺍﺮﺑ .ﺪﺷﺎﺑﻰﻣ ﻯﺎﻫپﻮﺗﻰﭘﺍﻒﻠﺘﺨﻣﻑﺩﺍﺮﺗﻪﺳﻯﻭﺎﺣﻰﭘﻮﺗﻰﭘﺍﺪﻨﭼﺪﻴﻤﺳﻼﭘﺶﺷ ﻰﺷﻮﻣپﻮﺗﻰﭘﺍﻚﻳ،HLA-A2ﻪﺑﻪﺘﺴﺑﺍﻭﻰﻧﺎﺴﻧﺍپﻮﺗﻰﭘﺍﻭﺩ) ﺭﻮﻛﺬﻣ ﻰﺣﺍﺮﻃ (ﻥﺎﺴﻧﺍﻭﺵﻮﻣﻦﻴﺑﻙﺮﺘﺸﻣپﻮﺗﻰﭘﺍﻚﻳﻭH2-Ddﻪﺑﻪﺘﺴﺑﺍﻭ ﻥﺎﻴﺑﻭﻩﺪﺷﻪﺘﺧﺎﺳﻰﻳﺎﻬﻨﺗﻪﺑﻭHBsAg-Sﻪﺑﻞﺼﺘﻣﺖﻟﺎﺣﻭﺩﺭﺩﻭ
ﺭﺍﺮﻗﻰﺑﺎﻳﺯﺭﺍﺩﺭﻮﻣﻰﻫﺎﮕﺸﻳﺎﻣﺯﺁﻒﻠﺘﺨﻣﻯﺎﻫﺵﻭﺭﺎﺑﺮﻈﻧﺩﺭﻮﻣﻥژ ﺭﻮﻛﺬﻣ ﻯﺎﻫﺪﻴﻤﺳﻼﭘﻰﻳﺍﺭﺎﻛ ﺯﺍﻥﺎﻨﻴﻤﻃﺍ ﺭﻮﻈﻨﻣ ﻪﺑ ﻦﻴﻨﭽﻤﻫ .ﺖﻓﺮﮔ ،ﻪﺘﺨﻳﺭﺍﺮﺗ ﺵﻮﻣ ﺭﺩ ﻰﻜﻳژﻮﻟﻮﻧﻮﻤﻳﺍ ﻖﻴﻗﺩ ﻯﺎﻫﺰﻴﻟﺎﻧﺁ ﻡﺎﺠﻧﺍ ﺯﺍ ﺶﻴﭘ ﺭﺩ،ﻩﺪﺷﻩﺪﻧﺎﺠﻨﮔﻰﺷﻮﻣﻯﺎﻫپﻮﺗﻰﭘﺍﺱﺎﺳﺍﺮﺑﺎﻬﻧﺁﻪﻴﻟﻭﺍﻰﻳﺍﺯﻰﻨﻤﻳﺍ .ﺪﺷﻰﺳﺭﺮﺑBALB/cﺵﻮﻣ
ﺎﻫ
ﺵﻭﺭ
ﻭ
ﺩﺍﻮﻣ
ﻰﭘﻮﺗﻰﭘﺍﺪﻨﭼﻯﺎﻫﺪﻴﻤﺳﻼﭘ
ﻪﺑﻪﺟﻮﺗﺎﺑHCVﺱﻭﺮﻳﻭCTLﻯﺎﻫپﻮﺗﻰﭘﺍﻰﺑﺎﻳﺯﺭﺍﻭﻰﻳﺎﺳﺎﻨﺷ
ﻯﺎﻫﭗﻴﺗﻮﻧژﺭﺩﺎﻬﻧﺁﻰﻟﺍﻮﺗﻪﺑﺎﺸﺗ،HLAﻝﻮﻜﻟﻮﻣﻪﺑﺎﻬﻧﺁﻝﺎﺼﺗﺍﻰﮔﮋﻳﻭ
ﻦﻳﺍﻯﺍﺮﺑ .ﺖﻓﺮﮔﺕﺭﻮﺻﺱﻭﺮﻳﻭﻯﺯﺎﺳﻙﺎﭘﺎﺑﺎﻬﻧﺁﻁﺎﺒﺗﺭﺍﻭﻒﻠﺘﺨﻣ ﺭﺩﻪﻛHCV ﻒﻠﺘﺨﻣﭗﻴﺗﻮﻧژ 18 ﻪﺑﻖﻠﻌﺘﻣﻪﻧﻮﻤﻧ 161 ﻥﺎﻴﻣﺯﺍﺭﻮﻈﻨﻣ
Los Alamos HCV Immunology Database ﻰﺗﺎﻋﻼﻃﺍ ﻩﺎﮕﻳﺎﭘ
ﻯﺎﻫﻥژﻰﺘﻧﺁ ﺯﺍ ﻪﺘﻓﺮﮔﺮﺑ CTL ﻯﺎﻫپﻮﺗﻰﭘﺍ ،ﺖﺳﺍ ﻩﺪﺷ ﺖﺒﺛ (34) ﻭﻪﺘﺷﺍﺩHLA-A2.1ﻯﺍﺮﺑﻯﺮﺗﻻﺎﺑﻝﺎﺼﺗﺍﻞﻴﻣﻪﻛNS3ﻭCore, E2
ﺱﻭﺮﻳﻭﻊﻳﺎﺷﭗﻴﺗﻮﻧژﻭﺩﻪﺑﺍﺭﺖﻫﺎﺒﺷﻦﻳﺮﺘﺸﻴﺑﺎﻬﻧﺁﻯﺍﻪﻨﻴﻣﺁﺪﻴﺳﺍﻰﻟﺍﻮﺗ ﺪﻌﺑﻡﺪﻗﺭﺩ .ﺪﻧﺪﺷﺏﺎﺨﺘﻧﺍﺮﺘﺸﻴﺑﻰﺑﺎﻳﺯﺭﺍﻯﺍﺮﺑ،ﺪﻨﺘﺷﺍﺩ1bﻭ1aﻰﻨﻌﻳ
ﻰﻨﻤﻳﺍﺦﺳﺎﭘﻰﻟﺎﻤﺘﺣﺍﻁﺎﺒﺗﺭﺍ ﺮﺑﻰﻨﺒﻣﺩﻮﺟﻮﻣﻱﺎﻫﺵﺭﺍﺰﮔﻪﺑ ﻪﺟﻮﺗﺎﺑ ﻰﻳﺎﻬﻧپﻮﺗﻰﭘﺍﻪﺳ،ﺖﻧﻮﻔﻋﻯﺯﺎﺳﻙﺎﭘﺎﺑﺎﻫپﻮﺗﻰﭘﺍﻪﻴﻠﻋ ﻩﺪﺷﺩﺎﺠﻳﺍ
H2-ﻪﺑﻪﺘﺴﺑﺍﻭﻪﻛ) E405پﻮﺗﻰﭘﺍﻩﻭﻼﻋﻪﺑN1406ﻭC132,E614
ﻥﺪﺷﻩﺪﻧﺎﺠﻨﮔﻯﺍﺮﺑ (ﺪﺷﺎﺑﻰﻣBALB/cﺵﻮﻣﺭﺩﻰﺑﺎﻳﺯﺭﺍﻞﺑﺎﻗﻭ Dd ﻝﻭﺪﺟ) ﺪﺷﻪﺘﻓﺮﮔﺮﻈﻧﺭﺩﻖﻴﻘﺤﺗ ﻦﻳﺍﻰﭘﻮﺗﻰﭘﺍ ﺪﻨﭼﻯﺎﻫﺭﺎﺘﺧﺎﺳﺭﺩ ﻯﺎﻫﻢﺘﻳﺭﻮﮕﻟﺍﺯﺍﻩﺩﺎﻔﺘﺳﺍﻭﻯﺮﺗﻮﻴﭙﻣﺎﻛﻯﺯﺎﺳﻪﻴﺒﺷﻚﻤﻛﺎﺑﺲﭙﺳ .(1
http://bio.dfci.harvard.edu/) ﻭHLA ﻪﺑ ﺪﻴﺘﭙﭘ ﻝﺎﺼﺗﺍ ﻰﻨﻴﺑﺶﻴﭘ
ﻪﺳ (http://paproc.de) ﻡﻭﺯﺎﺌﺗﻭﺮﭘ ﺵﺯﺍﺩﺮﭘ ﺰﻴﻧ ﻭ (/RANKPEP
ﺪﺷﻰﺣﺍﺮﻃﻯﺍﻪﻧﻮﮔﻪﺑﺭﻮﻛﺬﻣپﻮﺗﻰﭘﺍﺭﺎﻬﭼﺯﺍﻞﻜﺸﺘﻣپﻮﺗﻰﻠﭘﻰﻟﺍﻮﺗ ﺮﮕﻳﺩﻭﺩﻮﺷﺩﺎﺠﻳﺍﺎﻬﻧﺁﻞﺻﺎﻓﺪﺣﺭﺩﻰﻨﻴﺑﺎﻨﻴﺑﻯﺎﻫپﻮﺗﻰﭘﺍﻦﻳﺮﺘﻤﻛﻪﻛ ﻪﺠﻴﺘﻧﺭﺩ ﻭ ﻡﻭﺯﺎﺌﺗﻭﺮﭘ ﻂﺳﻮﺗ ﺎﻫپﻮﺗﻰﭘﺍ ﺵﺯﺍﺩﺮﭘﻯﺍﺮﺑ ﻂﻳﺍﺮﺷﻪﻜﻧﺁ ﻯﺮﻴﮔﺭﺍﺮﻗ ﻞﺤﻣﻰﻳﺎﺟﻪﺑﺎﺟﺎﺑ ﺎﻫﻰﮔﮋﻳﻭﻦﻳﺍ .ﺪﺷﺎﺑﻝﺁﻩﺪﻳﺍﺎﻬﻧﺁﻪﺿﺮﻋ ﻰﻧﺍﻭﺍﺮﻓﻝﻭﺪﺟﻪﺑﻪﺟﻮﺗﺎﺑﻡﺎﺠﻧﺍﺮﺳﻭﻩﺪﺷﻝﺮﺘﻨﻛﻢﻫﻪﺑﺖﺒﺴﻧﺎﻫپﻮﺗﻰﭘﺍ ﻪﺳ (/www.kazusa.org.jp/codon)ﻰﻧﺎﺴﻧﺍﻯﺎﻫﻥﻭﺪﻛﻯﺍﺮﺑtRNA
(35) SOEing PCRﺵﻭﺭﻪﺑﻭﻰﺣﺍﺮﻃﻪﻃﻮﺑﺮﻣﻯﺪﻴﺗﻮﺌﻠﻛﻮﻧﻰﻟﺍﻮﺗ
ﺪﻴﺗﻮﺌﻠﻛﻮﻧﻮﮕﻴﻟﺍﺖﻔﺟ ﻪﺳ،ﻰﻟﺍﻮﺗﻪﺳﻦﻳﺍﺖﺧﺎﺳ ﻯﺍﺮﺑ .ﺪﻧﺪﺷﻪﺘﺧﺎﺳ ﺎﺑ،ﺪﻨﺘﺷﺍﺩ(Overlap)ﻰﻧﺎﺷﻮﭘﻢﻫ 3/ﻪﻴﺣﺎﻧﺯﺎﺑ 21 ﺭﺩﻪﻛﻯﺯﺎﺑ 80-84
.ﺪﻧﺪﺷﻪﻴﻬﺗ (MWG Biotech Germany) ﻰﻧﺎﭙﻤﻛﺯﺍHPLCﺹﻮﻠﺧ
ﻑﺩﺍﺮﺗﺖﺧﺎﺳﻯﺍﺮﺑﻩﺪﺷﻰﺣﺍﺮﻃﻯﻮﮕﻴﻟﺍﺖﻔﺟﻚﻳﻪﻧﻮﻤﻧﻥﺍﻮﻨﻋﻪﺑ ﻰﻟﺍﻮﺗﺎﺑODNfﺯﺍﺪﻧﻮﺑﺕﺭﺎﺒﻋ(Polytope1) 1ﻩﺭﺎﻤﺷ
5′-ACCATGGGATCCGACCTGATGGGCTACATCCCTC TGGTGGGCGCCTCCGGCCCTTCCCAGAAGATCCA GCTGGTGCGGCT-3′
ﻰﻟﺍﻮﺗﺎﺑODNrﻭ 5′-ATGAATTCTTACACGGCGTTCAGGCCCAGGCCG GACAGCTTGATGGTGCAAGGGTAGTGCCACAGCC GCACCAGCTGGATCTTC-3′
ﻩﺩﺎﻔﺘﺳﺍﺎﺑ .ﺖﺳﺍﻩﺪﺷﺺﺨﺸﻣﺮﻳﺯﻂﺧﺎﺑﺎﻬﻧﺁﻥﺎﺷﻮﭘﻢﻫﻰﺣﺍﻮﻧﻪﻛ ﻝﺎﻤﻋﺍﻭ dNTP،ﻭ, Pfu DNA polymerase (Fermentas)Pﻢﻳﺰﻧﺁﺯﺍ
5 ﺕﺪﻣﻪﺑﻚﻳﺮﻫ) ﺩﺍﺮﮔﻰﺘﻧﺎﺳﻪﺟﺭﺩ 72 ﻭ 70 ،95 ﻰﺗﺭﺍﺮﺣﻞﻜﻴﺳﻚﻳ ﻰﺣﺍﻮﻧ،ﻥﺎﺷﻮﭘﻢﻫﻪﻴﺣﺎﻧﺭﺩﺎﻫﺪﻴﺗﻮﺌﻠﻛﻮﻧﻮﮕﻴﻟﺍﻥﺪﺷﺖﻔﺟﺯﺍﺲﭘ،(ﻪﻘﻴﻗﺩ ﻞﻣﺎﻛﻪﻌﻄﻗﺪﻌﺑﻪﻠﺣﺮﻣﺭﺩ .ﺪﺷﻪﺘﺧﺎﺳﻞﺑﺎﻘﻣﻪﺘﺷﺭﻯﻮﮕﻟﺍﻯﻭﺭﺯﺍﻰﻳﺎﻬﺘﻧﺍ ﻯﺎﻫﺮﻤﻳﺍﺮﭘﻥﺩﻭﺰﻓﺍﺎﺑPCRﺶﻨﻛﺍﻭﻚﻳﺭﺩﻮﮕﻟﺍﻥﺍﻮﻨﻋﻪﺑ
Pf (5′-TAGGATCCACCATGGACCTGATGGGCTAC-3′), Pr (5′
-ATGAATTCTTACACGGCG-3′)
،(ﮓﻧﺭﺮﭘﻰﺣﺍﻮﻧ)EcoRIﻭBamHIﻯﺎﻫﻢﻳﺰﻧﺁﺵﺮﺑﻰﻟﺍﻮﺗﻯﻭﺎﺣﻪﻛ ﺮﻴﺜﻜﺗ،ﺪﻧﺩﻮﺑ (Italicﻰﺣﺍﻮﻧ) Kozakﻰﻟﺍﻮﺗﻭﻪﻤﺟﺮﺗﻢﺘﺧﻭﻉﻭﺮﺷﻯﺎﻫﻥﻭﺪﻛ
ﻯﺎﻫپﻮﺗﻰﭘﺍﺯﺍﻒﻠﺘﺨﻣﺐﻴﺗﺮﺗﻪﺳﻪﻛﻩﺪﺷﺰﺘﻨﺳﻥژﻪﺳﺮﻫﺖﻳﺎﻬﻧﺭﺩ ﻰﺗﻮﻳﺭﺎﻛﻮﻳ ﻞﻗﺎﻧ ﺭﺩ ﻪﻧﺎﮔﺍﺪﺟ ﺭﻮﻃﻪﺑ ،ﺪﻧﺩﺮﻛﻰﻣ ﻥﺎﻴﺑ ﺍﺭ ﺚﺤﺑ ﺩﺭﻮﻣ ﻯﺎﻫﺪﻴﻤﺳﻼﭘﻥﺁﻞﺻﺎﺣﻪﻛﺪﻧﺪﺷﻥﻮﻠﻛ pcDNA3.1+(Invitrogen)
.(1 ﻞﻜﺷ) ﺪﻧﺩﻮﺑpcPOL3ﻭpcPOL2، pcPOL1ﻰﭘﻮﺗﻰﭘﺍﺪﻨﭼ
ﻥژﻪﺑﻞﺼﺘﻣﺮﻴﻏﻭﻞﺼﺘﻣﺕﺭﻮﺻﻪﺑCTLﻯﺎﻫپﻮﺗﻰﭘﺍﻒﻠﺘﺨﻣﻑﺩﺍﺮﺗﻪﺳﻩﺪﻨﻨﻛﺪﻛپﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﻤﺳﻼﭘﺖﺧﺎﺳﻞﺣﺍﺮﻣﺶﻳﺎﻤﻧ :1 ﻞﻜﺷ
ﻪﺑﺵﻭﺭﻦﻳﺍﻒﻠﺘﺨﻣﻑﺩﺍﺮﺗﻪﺳﺖﺧﺎﺳﻯﺍﺮﺑ .ﺖﺳﺍﻩﺪﺷﻩﺩﺍﺩﻥﺎﺸﻧﻑﺩﺍﺮﺗﻚﻳﻯﺍﺮﺑﺎﺠﻨﻳﺍﺭﺩSOEing PCRﻚﻴﺗﺎﻤﺷﻞﻜﺷ .HBsAg-S
ﻥﺎﺷﻮﭘﻢﻫﺪﻴﺗﻮﺌﻠﻛﻮﻧﻮﮕﻴﻟﺍﻭﺩSOEingﻪﻠﺣﺮﻣﺭﺩ .ﺪﺷﻪﺘﻓﺮﮔﺭﺎﻛﻪﺑﺕﻭﺎﻔﺘﻣﻯﺯﺎﺑ 80-84 ﻯﺎﻫﺪﻴﺗﻮﺌﻠﻛﻮﻧﻮﮕﻴﻟﺍﺎﺑﺭﺎﺑﻪﺳﻪﻧﺎﮔﺍﺪﺟﺕﺭﻮﺻ
ﺮﻴﺜﻜﺗﻞﺻﺎﺣﻪﻌﻄﻗPCRﻪﻠﺣﺮﻣﺭﺩ .ﺪﺷﻯﺯﺎﺳﺪﻨﻧﺎﻤﻫﻞﻤﻜﻣﻰﺣﺍﻮﻧﻭﻩﺩﺍﺩﺪﻳﺮﺒﻴﻫﻚﻳﻞﻴﻜﺸﺗﻰﺗﺭﺍﺮﺣﻞﻜﻴﺳﻝﺎﻤﻋﺍﻪﺠﻴﺘﻧﺭﺩODNrﻭODNf
ﺮﻴﺜﻜﺗHrﻭHfﺮﻤﻳﺍﺮﭘﻭﺩﻪﻠﻴﺳﻭﻪﺑﺰﻴﻧHBsAg-Sﻥژ .ﺪﻧﺪﺷﻪﻓﺎﺿﺍﻥﺁﻪﺑKozakﻰﻟﺍﻮﺗﻭﻢﺘﺧ،ﻉﻭﺮﺷﻯﺎﻫﻥﻭﺪﻛ،ﻢﻳﺰﻧﺁﺵﺮﺑﻰﻟﺍﻮﺗﻭﻩﺪﺷ
ﺭﺩﻭﻞﺼﺘﻣﺕﺭﻮﺻﻪﺑﺎﻳﻭﻰﺋﺎﻬﻨﺗﻪﺑپﻮﺗﻰﻠﭘﻯﺎﻫﻰﻟﺍﻮﺗﺖﻳﺎﻬﻧﺭﺩ .ﺪﺷﺩﺎﺠﻳﺍﻥﺁﻯﻭﺭﺮﺑﻰﻤﻳﺰﻧﺁﺵﺮﺑﻭﻪﻤﺟﺮﺗﻯﺍﺮﺑﻡﺯﻻﻯﺎﻫﻰﻟﺍﻮﺗﻭﻩﺪﺷ
،KozakﻰﻟﺍﻮﺗﻭﻪﻤﺟﺮﺗﻉﻭﺮﺷﻥﻭﺪﻛﻪﻧﺎﺸﻧﺐﻴﺗﺮﺗﻪﺑTAAﻭH،ATG/K .ﺪﻧﺪﺷﻥﻮﻠﻛpcDNA3.1ﺪﻴﻤﺳﻼﭘﺭﺩHBsAg-Sﻥژﺖﺋﺍﺮﻗﺐﻟﺎﻗ
ﻭN1406، E405، E614ﻯﺎﻫپﻮﺗﻰﭘﺍﻪﺑﻁﻮﺑﺮﻣﻰﻟﺍﻮﺗﻑﺮﻌﻣﺐﻴﺗﺮﺗﻪﺑﺰﻴﻧCﻭN ،E4 ،E6 .ﺪﻨﺘﺴﻫﻪﻤﺟﺮﺗﻢﺘﺧﻥﻭﺪﻛﻭHBsAg-Sﻥژ
.ﺪﻨﺷﺎﺑﻰﻣ C132
pcDNA3.1
5428 bp
ODNf
ODNr
HBsAg Overlap extension product
EcoRO Polytope Seq. (147bp)
H
1. SOEing step Pf
Hr
HindIII BamHI EcoRI
pcPOL3
pcHPOL3
pcHPOL1 pcPOL2
pcHPOL2 pcPOL1 K/ATG
H H H
E4
E4
E4
E4
E4 E4
TAA
TAA E6
E6 E6
E6
E6
E6 C
C
C
C
C
C
N N
N N
N K/ATG
K/ATG
K/ATG
K/ATG
K/ATG Hr
2. PCR step Pr
BamHI
HindIII BamHI
ﻰﭘﺍﺪﻨﭼﻯﺎﻫﻰﻟﺍﻮﺗﻰﻳﺍﺯﻰﻨﻤﻳﺍﺮﺑHBsAg-Sﺮﻴﺛﺎﺗﻰﺳﺭﺮﺑﺭﻮﻈﻨﻣﻪﺑ
ﻥژ،ﻰﭘﻮﺗ
HBsAg-S (GenBank accession X02496, REGION: 157-837)
ﺮﻤﻳﺍﺮﭘﻭﺩﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑ
Hf (5′-CCTAAGCTTACCATGGAGAACATCAC-3′)
Hr (5′-GCGGATCCTTAAATGTATACCCAAAGAC-3′) ﻰﺣﺍﻮﻧ) BamHI ﻭ HindIII ﻯﺎﻫﻢﻳﺰﻧﺁ ﺵﺮﺑ ﻰﻟﺍﻮﺗ ﻯﻭﺎﺣ ﻪﻛ ﻪﻤﺟﺮﺗ ﻢﺘﺧ ﻭ ﻉﻭﺮﺷ ،Kozak ﻯﺎﻫﻥﻭﺪﻛ ﺰﻴﻧ ﻭ (ﺮﻳﺯ ﻂﺧ ﻯﺍﺭﺍﺩ (ORF) ﺖﺋﺍﺮﻗ ﺐﻟﺎﻗ ﻚﻳﺭﺩ ﻭ ﻩﺪﺷﺮﻴﺜﻜﺗ ﺪﻧﺩﻮﺑ (Italic ﻰﺣﺍﻮﻧ) ﻥﻮﻠﻛpcDNA3.1+ ﺪﻴﻤﺳﻼﭘ ﺭﺩ ،پﻮﺗﻰﻠﭘ ﻰﻟﺍﻮﺗ ﻪﺳﺯﺍ ﻚﻳﺮﻫﺎﺑ
ﻭpcHPOL2 ،pcHPOL1 ﺮﮕﻳﺩ ﺪﻴﻤﺳﻼﭘﻪﺳﺐﻴﺗﺮﺗ ﻦﻳﺍﻪﺑ .ﺪﺷ پﻮﺗﻰﻠﭘ ﺪﻴﺘﭙﭘ ﻝﺎﺼﺗﺍ ﺯﺍ ﻞﻜﺸﺘﻣ ﺪﻳﺮﺒﻴﻫ ﻦﻴﺌﺗﻭﺮﭘ ﻪﻛ pcHPOL3
ﻪﺘﺧﺎﺳ ﺰﻴﻧ ﺪﻧﺩﺮﻛﻰﻣ ﻥﺎﻴﺑ ﺍﺭ HBsAg-S ﻞﻴﺴﻛﻮﺑﺮﻛ ﻯﺎﻬﺘﻧﺍ ﻪﺑ
ﺯﺍ ﻥﺎﻨﻴﻤﻃﺍ ﺯﺍ ﺲﭘ ﻕﻮﻓ ﺪﻴﻤﺳﻼﭘ ﺶﺷ ﻪﻴﻠﻛ .(1 ﻞﻜﺷ) ﺪﻧﺪﺷ ﺪﻴﻤﺳﻼﭘﺺﻴﻠﺨﺗﻯﺎﻫﻥﻮﺘﺳﺯﺍﻩﺩﺎﻔﺘﺳﺍ ﺎﺑﻯﺪﻴﺗﻮﺌﻠﻛﻮﻧ ﻰﻟﺍﻮﺗﺖﺤﺻ ﻑﺬﺣﺎﺑNucleoBond Giga (Macherey-Nagel, Germany)
.ﺪﻧﺪﺷﺺﻴﻠﺨﺗﺖﻴﻛﻞﻤﻌﻟﺍﺭﻮﺘﺳﺩﻖﺑﺎﻄﻣﻭﻦﻴﺴﻛﻮﺗﻭﺪﻧﺍ
ﺵﻮﮔﺮﺧﺯﺍﻝﺎﻧﻮﻠﻛﻰﻠﭘ ﻡﺮﺳﻰﺘﻧﺁﻪﻴﻬﺗ ﻪﺑﺍﺪﺘﺑﺍﻖﻴﻘﺤﺗﻦﻳﺍ پﻮﺗﻰﻠﭘ ﻯﺎﻫﺪﻴﺘﭙﭘﻪﻴﻠﻋﻯﺩﺎﺑﻰﺘﻧﺁﻪﻴﻬﺗﻥﺎﻜﻣﺍ
Bﻝﻮﻠﺳﻯﺎﻫپﻮﺗﻰﭘﺍﻰﻳﺎﺳﺎﻨﺷﻢﺘﻳﺭﻮﮕﻟﺍﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑﻭin silicoﺕﺭﻮﺻ
(http: //bio.dfci. harvard. edu/tools/antigenic. htmL)
ﻯﺩﺎﺑﻰﺘﻧﺁ ،(36) ﺩﺭﺍﺪﻧﺎﺘﺳﺍ ﺵﻭﺭﻚﻳ ﻝﺎﻤﻋﺍﺎﺑ ﺲﭙﺳ ﻩﺪﺷﻰﺳﺭﺮﺑ(l
.ﺪﺷﻪﻴﻬﺗﻪﻧﺎﮔﺍﺪﺟﺭﻮﻃﻪﺑﺎﻫﺪﻴﺘﭙﭘﺯﺍﻚﻳﺮﻫﻪﻴﻠﻋﻰﺷﻮﮔﺮﺧﻝﺎﻧﻮﻠﻛﻰﻠﭘ
ﻡﺮﮔﻰﻠﻴﻣ 1 ﻞﻣﺎﺷﻥژﻮﻧﻮﻤﻳﺍ ﻁﻮﻠﺨﻣ ﺯﺍﺮﺘﻴﻟﻰﻠﻴﻣ 1 ﻪﺻﻼﺧﺭﻮﻃ ﻪﺑ
ﻩﺎﮕﺸﻧﺍﺩ ﻰﻤﻴﺷﻮﻴﺑ ﻮﺘﻴﺘﺴﻧﺍ ﻂﺳﻮﺗ ﻩﺪﺷ ﺰﺘﻨﺳ) ﻚﻴﺘﺘﻨﺳ ﺪﻴﺘﭙﭘ ﺯﺍ ﺪﻧﻭﺮﻓ ﺺﻗﺎﻧ ﻥﺍﻮﺟﺍ ﻯﻭﺎﺴﻣ ﻢﺠﺣ ﺎﺑ ﻩﺪﺷ ﻁﻮﻠﺨﻣ (ﺲﻴﺋﻮﺳ ﻥﺍﺯﻮﻟ ﻪﺑ ﻯﺪﻠﺟﺮﻳﺯ ﺕﺭﻮﺻ ﻪﺑ (Incomplete Freund’s Adjuvant)
،ﻩﺎﻣﻚﻳﺯﺍﺲﭘﺭﻭﺁﺩﺎﻳﻖﻳﺭﺰﺗﻝﺎﺒﻧﺩﻪﺑ .ﺪﺷﻖﻳﺭﺰﺗﻪﻫﺎﻣ 3 ﺵﻮﮔﺮﺧ ﻦﻳﺮﺘﺸﻴﺑﻦﺘﻓﺎﻳﺖﻬﺟﻒﻠﺘﺨﻣﻰﻧﺎﻣﺯﻞﺻﺍﻮﻓﺭﺩﺵﻮﮔﺮﺧﻥﻮﺧﻪﻧﻮﻤﻧ .ﺖﻓﺮﮔﺭﺍﺮﻗﻰﺳﺭﺮﺑﺩﺭﻮﻣﻢﻴﻘﺘﺴﻣﺮﻴﻏﻯﺍﺰﻳﻻﺍﺵﻭﺭﺎﺑﻯﺩﺎﺑﻰﺘﻧﺁﺭﺎﻴﻋ
ﻰﻟﻮﻠﺳﺕﺍﺰﻴﻟﻪﻴﻬﺗﻭﺖﻜﻓﺲﻧﺍﺮﺗ
ﻥﻮﻤﻴﻣﻪﻴﻠﻛﺯﺍﻥﺁﺄﺸﻨﻣﻪﻛCos-7ﻰﻟﻮﻠﺳﻩﺩﺭﻯﺭﺍﺪﻬﮕﻧﻭﺖﺸﻛ
ﺭﻻﻮﻣﻰﻠﻴﻣ 2 ﻯﻭﺎﺣ DMEMﺖﺸﻛﻂﻴﺤﻣﺭﺩ،ﺪﺷﺎﺑﻰﻣﻰﻳﺎﻘﻳﺮﻓﺁﺰﺒﺳ
ﻡﺮﮔﻰﻠﻴﻣ 100 ،ﻦﻴﻠﻴﺳﻰﻨﭘ ﺮﺘﻴﻟﻰﻠﻴﻣ ﺭﺩ ﺪﺣﺍﻭ 100 ،ﻦﻴﻣﺎﺗﻮﻠﮔ-ﻝﺍ
،(FCS) ﻭﺎﮔ ﻦﻴﻨﺟ ﻡﺮﺳ ﺪﺻﺭﺩ 10 ﻭ ﻦﻴﺴﻳﺎﻣﻮﺘﭘﺮﺘﺳﺍ ﺮﺘﻴﻟﻰﻠﻴﻣ ﺭﺩ
ﺕﺭﻮﺻ CO2 ﺪﺻﺭﺩ 5 ﺮﻔﺴﻤﺗﺍ ﻭ ﺩﺍﺮﮔﻰﺘﻧﺎﺳ ﻪﺟﺭﺩ 37 ﻯﺎﻣﺩ ﺭﺩ
ﻯﺭﺎﺠﺗ ﺖﻴﻛ ﺯﺍ ﻩﺩﺎﻔﺘﺳﺍﺎﺑ ﻭ ﻦﺸﻜﻓﻮﭙﻴﻟ ﺵﻭﺭ ﻪﺑ ﻝﻮﻠﺳﻦﻳﺍ .ﺖﻓﺮﮔ ﻯﺎﻫﺪﻴﻤﺳﻼﭘﺎﺑ (Transient) ﺖﻗﻮﻣﺕﺭﻮﺻﻪﺑ PolyFect (Qiagen) ﺖﻜﻓﺲﻧﺍﺮﺗ ﺲﻜﻠﭙﻤﻛ ﺭﻮﻈﻨﻣ ﻦﻳﺍ ﻯﺍﺮﺑ .ﺪﺷ ﺖﻜﻓﺲﻧﺍﺮﺗ پﻮﺗﻰﻠﭘ ﻞﻤﻌﻟﺍﺭﻮﺘﺳﺩﻖﺒﻃﺖﻜﻓﻰﻠﭘﻝﻮﻠﺤﻣﻭﺪﻴﻤﺳﻼﭘﺐﺳﺎﻨﻣﺮﻳﺩﺎﻘﻣﺯﺍﻞﻜﺸﺘﻣ
ﺖﻋﺎﺳ 72 .ﺪﺷﻪﻓﺎﺿﺍﻝﻮﻠﺳﺖﺸﻛﻪﺑﻰﻤﺘﻳﺭﺎﮕﻟﺯﺎﻓﺭﺩﻭﻩﺪﺷﻪﻴﻬﺗﺖﻴﻛ
ﺖﺤﺗﺎﻫﻝﻮﻠﺳ،ﻰﻳﻭﺭﺖﺸﻛﻂﻴﺤﻣﻯﺭﺍﺪﻬﮕﻧﻭﻯﺭﻭﺁﻊﻤﺟﻦﻤﺿﺪﻌﺑ ﺕﺍﺰﻴﻟ ﻪﻴﻬﺗ ﺭﻮﻈﻨﻣﻪﺑ ﺎﻳ ﻭ ﻪﺘﻓﺮﮔﺭﺍﺮﻗ ﺲﻧﺎﺳﺭﻮﻠﻓﻮﻧﻮﻤﻳﺍ ﻯﺰﻴﻣﺁﮓﻧﺭ ﺮﺑﻪﻴﻧﺎﺛ 30 ﺕﺪﻣﻪﺑﺭﺎﺑﺮﻫﻭﻪﻘﻴﻗﺩ 1 ﻰﻧﺎﻣﺯﻞﺻﺍﻮﻓﺎﺑﺭﺎﺑﺞﻨﭘ،ﻰﻟﻮﻠﺳ ﻪﺑ ﻞﺻﺎﺣ ﺕﺍﺰﻴﻟ .ﺪﻨﺘﻓﺮﮔ ﺭﺍﺮﻗﻥﻮﻴﺳﺎﻜﻴﻧﻮﺳﺍﺮﺘﻟﻭﺍ ﺮﺛﺍ ﺖﺤﺗ ﺦﻳﻯﻭﺭ ﺰﻴﻟﺎﻧﺁﺖﻬﺟﻰﻳﻭﺭﻊﻳﺎﻣﻭﻩﺪﺷژﻮﻴﻔﻳﺮﺘﻧﺎﺳg2000 ﺭﺩﻪﻘﻴﻗﺩ 20 ﺕﺪﻣ ﻪﻌﻄﻗﺪﻗﺎﻓﻯﺎﻫﺪﻴﻤﺳﻼﭘﺎﺑﻩﺪﺷﺖﻜﻓﺲﻧﺍﺮﺗﻯﺎﻫﻝﻮﻠﺳ .ﺪﺷﻯﺭﻭﺁﻊﻤﺟ
(HBsAg ﻥژﻱﻭﺎﺣ pcDNA)ﻭ pcHﻭ pcDNA3.1ﻰﻨﻌﻳپﻮﺗﻰﻠﭘ
ﺮﻈﻧﺭﺩﻰﻔﻨﻣﻝﺮﺘﻨﻛﻥﺍﻮﻨﻋﻪﺑﻩﺪﺸﻧﺖﻜﻓﺲﻧﺍﺮﺗCos-7ﻯﺎﻫﻝﻮﻠﺳﺰﻴﻧﻭ
.ﺪﻧﺪﺷﻪﺘﻓﺮﮔ
پﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﺘﭙﭘﻥﺎﻴﺑﻰﺳﺭﺮﺑ ﺯﺍﺲﭘﻰﻟﻮﻠﺳﺕﺍﺰﻴﻟﺭﺩﺩﻮﺟﻮﻣﻯﺎﻫﻦﻴﺌﺗﻭﺮﭘ :ﺕﻼﺑﻥﺮﺘﺳﻭ .ﻒﻟﺍ ﺵﻭﺭﻚﻳﺱﺎﺳﺍﺮﺑSDS-PAGEﺪﺻﺭﺩ 15 ﻝژﻯﻭﺭﺮﺑﻯﺯﺎﺳﺍﺪﺟ ﻦﻴﺌﺗﻭﺮﭘﻪﺑﻁﻮﺑﺮﻣﺪﻧﺎﺑﻭﻩﺪﺷﻞﻘﺘﻨﻣﺰﻟﻮﻠﺳﻭﺮﺘﻴﻧﺎﺸﻏﻪﺑ (37) ﺩﺭﺍﺪﻧﺎﺘﺳﺍ ﺪﺿ ﻝﺎﻧﻮﻠﻛﻮﻧﻮﻣ ﻱﺩﺎﺑ ﻲﺘﻧﺁ ﺯﺍ ﻩﺩﺎﻔﺘﺳﺍ ﺎﺑ HBs-polytope ﺪﻳﺮﺒﻴﻫ HRPﺎﺑﻪﮔﻭﮋﻨﻛﻪﻳﻮﻧﺎﺛﻯﺩﺎﺑﻰﺘﻧﺁﻭHBsAg (Cedarlane, Canada) .ﺪﺷﺖﻳﻭﺭﻞﺑﺎﻗDABﻯﺍﺰﮕﻧﺭﻝﻮﻠﺤﻣﻥﺩﻭﺰﻓﺍﻭ(Sigma, USA)ﻭ
ﻯﻭﺭ ﺮﺑ ﻩﺪﺷ ﻩﺩﺍﺩ ﺖﺸﻛ ﻯﺎﻫﻝﻮﻠﺳ :ﺲﻧﺎﺳﺭﻮﺋﻮﻠﻓﻮﻧﻮﻤﻳﺍ .ﺏ ﺲﭘﺖﻋﺎﺳ 72 ،(Glass Cover Slips) ﻯﺍﻪﺸﻴﺷﺹﻮﺼﺨﻣﻯﺎﻫﻞﻣﻻ ﻪﻘﻴﻗﺩ 20 ﺕﺪﻣﻪﺑﺪﺻﺭﺩ4 ﺪﻴﺋﺪﻟﺎﻣﺭﻮﻓﺍﺭﺎﭘﺕﺭﻭﺎﺠﻣﺭﺩﺖﻜﻓﺲﻧﺍﺮﺗﺯﺍ
HCVﻪﻴﻠﻋﻰﭘﻮﺗﻰﭘﺍﺪﻨﭼﻦﺴﻛﺍﻭﻰﺣﺍﺮﻃﺭﺩﻪﺘﻓﺭﺭﺎﻛﻪﺑCTLﻯﺎﻫپﻮﺗﻰﭘﺍ :1 ﻝﻭﺪﺟ
ﻊﺟﺮﻣ
ﻝﺎﺼﺗﺍﻰﺘﻴﻨﻴﻓﺍ
MHCﻪﺑ
(%Score)$
ﺶﺷﻮﭘﺪﺻﺭﺩ
ﭗﻴﺗﻮﻧژ
ﻪﺑﻰﮕﺘﺴﺑﺍﻭ
MHC #پﻮﺗﻰﭘﺍﻞﺤﻣ
ﻪﻳﻮﺳﺭﺩ
HCV-H
ﻊﺟﺮﻣﻥژﻰﺘﻧﺁ
ﻯﺪﻴﺳﺍﻮﻨﻴﻣﺁﻰﻟﺍﻮﺗ
پﻮﺗﻰﭘﺍ
1b‡ 1a†
(45-47) 66/7-63/6
96/1 100
A2.1/H-2d 132-142
Core DLMGYIPLVGA*
C132
(47،44)
50 2 75
A2.1
614-622
E2 RLWHYPCTI
E614
(49،47)
58/5 25/2 0
A2.1
1406-1415
NS3 KLSGLGLNAV
N1406
(48) 52/9
2/9 0
H-2d 405-414
E2 SGPSQKIQLV
E405
.ﺖﺳﺍﻩﺪﺷﻩﺩﺍﺩﻥﺎﺸﻧ(FDA) HCV-Hﻂﺳﻮﺗﻩﺪﺷﺍﺪﺟﻪﻳﻮﺳﻰﻟﺍﻮﺗﺱﺎﺳﺍﺮﺑﺎﻫﻦﻴﺌﺗﻭﺮﭘﻞﺤﻣ #
.1aﭗﻴﺗﻮﻧژﻪﺑﻖﻠﻌﺘﻣﻩﺪﺷﻰﺳﺭﺮﺑﻪﻧﻮﻤﻧ 8 ﺭﺩﻯﺪﻴﺳﺍﻮﻨﻴﻣﺁﻰﻟﺍﻮﺗﻰﻧﺎﺴﻜﻳ †
.1bﭗﻴﺗﻮﻧژﻪﺑﻖﻠﻌﺘﻣﻩﺪﺷﻰﺳﺭﺮﺑﻪﻧﻮﻤﻧ 103 ﺭﺩﻯﺪﻴﺳﺍﻮﻨﻴﻣﺁﻰﻟﺍﻮﺗﻰﻧﺎﺴﻜﻳ ‡
ﻪﺒﺳﺎﺤﻣﺭﺍﺰﻓﺍﻡﺮﻧﻂﺳﻮﺗ (ﺩﺭﺍﺩﺍﺭﻰﺘﻴﻨﻴﻓﺍﻦﻳﺮﺘﺸﻴﺑﻩﺪﺷﺺﺨﺸﻣMHCﻯﺍﺮﺑﻪﻛ) consensusﻰﻟﺍﻮﺗﻪﺑپﻮﺗﻰﭘﺍﻰﻟﺍﻮﺗﻰﻜﻳﺩﺰﻧﻪﺑﻪﺟﻮﺗﺎﺑﻩﺪﺷﺮﻛﺫﺪﺻﺭﺩ $
.ﺖﺳﺍﻩﺪﺷ
(PBS) ﺕﺎﻔﺴﻓﺮﻓﺎﺑﺭﺩﻮﺷﻭﺖﺴﺷﺯﺍﺲﭘ .ﺪﻧﺪﺷﺲﻜﻴﻓﻕﺎﺗﺍﻯﺎﻣﺩﺭﺩ ﺭﺩﻪﻘﻴﻗﺩ 10 ﺕﺪﻣﻪﺑﺎﻫﻞﻣﻻ،ﻪﻨﻴﻣﺯﺲﻧﺎﺳﺭﻮﺋﻮﻠﻓﺯﺍﻦﺘﺳﺎﻛﺭﻮﻈﻨﻣﻪﺑ ﻯﻮﺷﻭﺖﺴﺷﻝﺎﺒﻧﺩﻪﺑﻭﻪﺘﻓﺮﮔﺭﺍﺮﻗﻡﻮﻴﻧﻮﻣﺁﺭﻭﺮﻠﻛﺭﻻﻮﻣﻰﻠﻴﻣ 50 ﻝﻮﻠﺤﻣ ﺫﻮﻔﻧ X-100ﻥﻮﺘﻳﺮﺗﺪﺻﺭﺩ 0/1 ﻝﻮﻠﺤﻣﻂﺳﻮﺗ ﺍﺪﺘﺑﺍﺎﻫﻝﻮﻠﺳ،ﺩﺪﺠﻣ ﺕﺭﻭﺎﺠﻣﺭﺩﺐﻴﺗﺮﺗﻪﺑﻪﻘﻴﻗﺩ 45 ﺕﺪﻣﻪﺑﻮﺷﻭﺖﺴﺷﺯﺍﺲﭘﻭﻩﺪﺷﺮﻳﺬﭘ
FITCﺎﺑﻪﮔﻭﮋﻨﻛﻪﻳﻮﻧﺎﺛﻯﺩﺎﺑﻰﺘﻧﺁﺲﭙﺳﻭHBsAgﺪﺿﻯﺩﺎﺑﻰﺘﻧﺁﺎﺑ
ﺯﺍﺪﻌﺑﻭﻞﺒﻗ .ﺪﻧﺪﺷﻪﺑﻮﻜﻧﺍ ﺩﺍﺮﮔﻰﺘﻧﺎﺳﻪﺟﺭﺩ 4 ﻯﺎﻣﺩﺭﺩ (Sigma)a
ﺖﻳﺎﻬﻧﺭﺩﻭﺪﻧﺪﺷﻪﺘﺴﺷPBSﺮﻓﺎﺑﻂﺳﻮﺗﺎﻫﻝﻮﻠﺳﻯﺩﺎﺑﻰﺘﻧﺁﺮﻫﻥﺩﻭﺰﻓﺍ
ﻯﺎﻫﻝﻮﻠﺳﺎﺑﻪﺴﻳﺎﻘﻣﺭﺩﻩﺪﺷﺖﻜﻓﺲﻧﺍﺮﺗﻯﺎﻫﻝﻮﻠﺳﺲﻧﺎﺳﺭﻮﺋﻮﻠﻓﻥﺍﺰﻴﻣ پﻮﻜﺳﻭﺮﻜﻴﻣ ﻂﺳﻮﺗ (ﻰﻔﻨﻣ ﻝﺮﺘﻨﻛ ﻥﺍﻮﻨﻋ ﻪﺑ) ﻩﺪﺸﻧ ﺖﻜﻓﺲﻧﺍﺮﺗ .ﺖﻓﺮﮔﺭﺍﺮﻗﻰﺳﺭﺮﺑﺩﺭﻮﻣZeiss Axioskopﺲﻧﺎﺳﺭﻮﺋﻮﻠﻓ
ﻯﺭﺍﺬﮔﻪﻜﻟﻯﺎﻫﻪﻧﻮﻤﻧﺭﺎﺸﺘﻧﺍﻥﺍﺰﻴﻣﺯﺍﻦﺘﺳﺎﻛﺭﻮﻈﻨﻣﻪﺑ :ﺕﻼﺑﺕﺍﺩ .ﺝ ﺕﺎﻌﻄﻗﻩﺯﺍﺪﻧﺍﻪﺑﺍﺪﺘﺑﺍﺯﻮﻟﻮﻠﺳﻭﺮﺘﻴﻧﺎﺸﻏ،ﺖﺳﺩﻚﻳﻁﺎﻘﻧﻦﺘﺷﺍﺩﻭﻩﺪﺷ ﻚﻳﻯﺎﻫﻚﻫﺎﭼﺭﺩﺕﺎﻌﻄﻗﻥﺩﺍﺩﺭﺍﺮﻗﺯﺍﺲﭘﻭﻩﺪﺷﭻﻧﺎﭘﻯﺮﺘﻣﻰﻠﻴﻣ 4 ﺩﺭﺍﺪﻧﺎﺘﺳﺍﺵﻭﺭﻖﺒﻃﻚﻴﺘﺘﻨﺳﻯﺎﻫﺪﻴﺘﭙﭘﺎﻳﻭ ﻰﻟﻮﻠﺳﺕﺍﺰﻴﻟ ،ﺍﺰﻳﻻﺍﺖﻴﻠﭘ ﻯﺎﻣﺩﺭﺩﺖﻋﺎﺳ 1 ﺕﺪﻣﻪﺑﺖﻴﻠﭘ .ﺪﻧﺪﺷﻯﺭﺍﺬﮔﻪﻧﻮﻤﻧﺎﺸﻏﻯﻭﺭﺮﺑ (38) ﻂﺳﻮﺗﺍﺪﺘﺑﺍﺎﻫﻚﻫﺎﭼﻥﻭﺭﺩﺕﺎﻌﻄﻗﻭﻩﺪﺷﻪﺑﻮﻜﻧﺍﺩﺍﺮﮔﻰﺘﻧﺎﺳﻪﺟﺭﺩ 37
ﺕﺪﻣ ﻪﺑﺲﭙﺳ ﻭﻩﺪﺷ ﻙﻼﺑﺪﺻﺭﺩ 3 (BSA) ﻯﻭﺎﮔﻡﺮﺳﻦﻴﻣﻮﺒﻟﺁ
ﻭ پﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﺘﭙﭘﻪﻴﻠﻋﻩﺪﺷﻪﻴﻬﺗﻰﺷﻮﮔﺮﺧﻡﺮﺳﻰﺘﻧﺁﺎﺑﻪﻘﻴﻗﺩ 30 ﺯﺍﺲﭘ .ﺪﻧﺪﺷﺭﻭﺎﺠﻣHRPﺎﺑﻪﮔﻭﮋﻨﻛﻪﻳﻮﻧﺎﺛﻯﺩﺎﺑﻰﺘﻧﺁﺎﺑﻥﺁﻝﺎﺒﻧﺩﻪﺑ
ﻚﻳﻯﻭﺭﺮﺑﺍﺭﻩﺪﺷﭻﻧﺎﭘﺕﺎﻌﻄﻗDABﻝﻮﻠﺤﻣﻪﻠﻴﺳﻭﻪﺑﺎﻫﻪﻜﻟﺭﻮﻬﻇ
.ﺪﻧﺪﺷﻯﺭﺍﺩﺮﺑﺲﻜﻋﻝﺎﺘﻴﺠﻳﺩﻦﻴﺑﺭﻭﺩﻂﺳﻮﺗﻭﻩﺩﺍﺩﺭﺍﺮﻗﺪﻴﻔﺳﻪﺤﻔﺻ ﻢﻴﻈﻨﺗپﺎﺷﻮﺗﻮﻓﺭﺍﺰﻓﺍﻡﺮﻧﺎﺑﻞﺻﺎﺣﺲﻜﻋﺖﻴﻓﺎﻔﺷﻭﺡﻮﺿﻭﺖﻳﺎﻬﻧﺭﺩ ﻩﺪﺸﻧﺖﻜﻓﺲﻧﺍﺮﺗﻯﺎﻫﻝﻮﻠﺳﺕﺍﺰﻴﻟﻭپﻮﺗﻰﻠﭘﻚﻴﺘﺘﻨﺳﻯﺎﻫﺪﻴﺘﭙﭘ .ﺪﺷ .ﺪﻧﺪﺷﻩﺩﺎﻔﺘﺳﺍﻰﻔﻨﻣﻭﺖﺒﺜﻣﻝﺮﺘﻨﻛﻥﺍﻮﻨﻋﻪﺑﺐﻴﺗﺮﺗﻪﺑ ﻝﻮﻠﺤﻣ ﻪﻠﻴﺳﻭﻪﺑ ﻰﻟﻮﻠﺳ RNA ﺺﻴﻠﺨﺗﻝﺎﺒﻧﺩ ﻪﺑ :RT-PCR .ﺩ ﻩﺩﺎﻔﺘﺳﺍﺎﺑ ﻪﻃﻮﺑﺮﻣ cDNAﺍﺪﺘﺑﺍInvitrogen ﺖﺧﺎﺳ Trizol ﻱﺭﺎﺠﺗ Reverse Transcriptase ﻢﻳﺰﻧﺁﻭRandom Hexamerﺮﻤﻳﺍﺮﭘﺯﺍ DNA ﺎﺑ RNA ﻰﮔﺩﻮﻟﺁ ﻡﺪﻋ ﺯﺍ ﻥﺎﻨﻴﻤﻃﺍ ﺖﻬﺟ .(39) ﺪﺷﻪﺘﺧﺎﺳ ﺲﭙﺳ .ﺪﺷﻪﻴﻬﺗﺰﻴﻧRTﻢﻳﺰﻧﺁﻥﺩﻭﺰﻓﺍﻥﻭﺪﺑﻝﺮﺘﻨﻛﻪﻧﻮﻤﻧﻯﺪﻴﻤﺳﻼﭘ
ﺭﺎﻨﻛ ﺭﺩﻝﺮﺘﻨﻛ ﻭﺖﺴﺗﻪﻟﻮﻟ ﻭﺩﺯﺍ ﻚﻳﺮﻫﻝﻮﺼﺤﻣﺯﺍﺮﺘﻴﻟﻭﺮﻜﻴﻣ 2 ﻪﻌﻄﻗﻥﻭﺪﺑﺪﻴﻤﺳﻼﭘﺎﺑﻩﺪﺷﺖﻜﻓﺲﻧﺍﺮﺗﻯﺎﻫﻝﻮﻠﺳﻪﺑﻁﻮﺑﺮﻣcDNA
ﺯﺍﻝﻮﻣﻮﻜﻴﭘ 20 ﻯﻭﺎﺣPCRﺩﺭﺍﺪﻧﺎﺘﺳﺍﺶﻨﻛﺍﻭﻚﻳﺭﺩ (pcDNA3.1) ﻰﺗﺭﺍﺮﺣﻪﻣﺎﻧﺮﺑﻞﻜﻴﺳ 27 ﻖﺒﻃپﻮﺗﻰﻠﭘﻯﺎﻫﻥژﻰﺻﺎﺼﺘﺧﺍﻯﺎﻫﺮﻤﻳﺍﺮﭘ .ﺪﻳﺩﺮﮔﺮﻴﺜﻜﺗ (ﺩﺍﺮﮔﻰﺘﻧﺎﺳﻪﺟﺭﺩ 72 ﻪﻴﻧﺎﺛ 30 ﻭ 50 ﻪﻴﻧﺎﺛ 30 ،95 ﻪﻴﻧﺎﺛ 20) ﻪﻠﻴﺳﻭﻪﺑﺯﺭﺎﮔﺁﺪﺻﺭﺩ 2 ﻝژﻯﻭﺭﺮﺑﺯﺭﻮﻓﻭﺮﺘﻜﻟﺍﺯﺍﺲﭘPCRﻝﻮﺼﺤﻣ
.ﺪﺷﻩﺪﻫﺎﺸﻣ UVP BioDoc-It (Cambridge, Uk) ﻢﺘﺴﻴﺳ
-HBs-poly ﺕﺍﺭﺫ ﺢﺷﺮﺗ ﻥﺍﺰﻴﻣ ﻰﺳﺭﺮﺑ ﺭﻮﻈﻨﻣ ﻪﺑ :ﺍﺰﻳﻻﺍ (ﻩ ﻰﻳﻭﺭ ﺖﺸﻛ ﻂﻴﺤﻣ ﺎﻫﻝﻮﻠﺳ ﺖﻜﻓﺲﻧﺍﺮﺗ ﺯﺍ ﺲﭘ ﺖﻋﺎﺳ 72 ،tope
ﻩﺪﻨﻨﻛﻆﻴﻠﻐﺗ ﻯﺎﻫﻪﻟﻮﻟ ﻂﺳﻮﺗ ﺮﺑﺍﺮﺑ ﻩﺩ ﻥﺍﺰﻴﻣ ﺎﺗ ﻭ ﻩﺪﺷ ﻯﺭﻭﺁﻊﻤﺟ ﺎﺑ ﺲﭙﺳ .ﺪﻧﺪﺷﻆﻴﻠﻐﺗ Vivaspin 2, 3000 MWCO (Sartorius) -Hepanos)ﺍﺰﻳﻻﺍﺵﻭﺭﻪﺑHBsAgﻯﺮﻴﮔﻩﺯﺍﺪﻧﺍﻯﺭﺎﺠﺗﺖﻴﻛﺯﺍﻩﺩﺎﻔﺘﺳﺍ
ﻂﻴﺤﻣﺭﺩﻩﺪﺷﺢﺷﺮﺗﺪﻳﺮﺒﻴﻫﺕﺍﺭﺫﺭﺍﺪﻘﻣ(tika, Biomérieux, France
ﻥﺍﺰﻴﻣﻯﺯﺎﺳﺖﺧﺍﻮﻨﻜﻳﺖﻬﺟﻪﻛﺖﺳﺍﺮﻛﺫﻪﺑﻡﺯﻻ .ﺪﺷﻯﺮﻴﮔﻩﺯﺍﺪﻧﺍﺖﺸﻛ
-pcDﺪﻴﻤﺳﻼﭘﻥﺎﻣﺯﻢﻫﺖﻜﻓﺲﻧﺍﺮﺗﺯﺍپﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﻤﺳﻼﭘﺖﻜﻓﺲﻧﺍﺮﺗ
ﻩﺩﺎﻔﺘﺳﺍﻰﻟﻮﻠﺳﺕﺍﺰﻴﻟﺭﺩﺯﺍﺪﻳﺯﻮﺘﻛﻻﺎﮔﺎﺘﺑﻢﻳﺰﻧﺁﺶﺠﻨﺳﻭNA3.1/LacZ ﻥﺍﺰﻴﻣﺎﺑﻪﺴﻳﺎﻘﻣﺭﺩﻰﺘﺧﺍﻮﻨﻜﻳﺐﻳﺍﺮﺿﻝﺎﻤﻋﺍﺯﺍﺲﭘﺍﺰﻳﻻﺍﺞﻳﺎﺘﻧﻭ (40) ﻩﺪﺷ ﻯﺎﻫﻝﻮﻠﺳﺯﺍ (Wild Type) ﻩﺪﺸﻧﻯﺭﺎﻜﺘﺳﺩHBsAg-Sﺕﺍﺭﺫﺢﺷﺮﺗ
.ﺪﻧﺪﺷﻥﺎﻴﺑﺪﺻﺭﺩﺕﺭﻮﺻﻪﺑ،pcHﺪﻴﻤﺳﻼﭘﺎﺑﻪﺘﻜﻓﺲﻧﺍﺮﺗ
ﺎﻫﺵﻮﻣﻯﺯﺎﺳﻦﻤﻳﺍ
ﺵﺭﻭﺮﭘﺰﻛﺮﻣﺯﺍﻦﺳﻪﺘﻔﻫ 6-8 ﺎﺑﻩﺩﺎﻣBALB/c (H-2d) ﻯﺎﻫﺵﻮﻣ ﻭﻪﻴﻬﺗﺝﺮﻛﺭﺩﻥﺍﺮﻳﺍﺭﻮﺘﺳﺎﭘﻮﺘﻴﺘﺴﻧﺍﻰﻫﺎﮕﺸﻳﺎﻣﺯﺁﺕﺎﻧﺍﻮﻴﺣﻯﺭﺍﺪﻬﮕﻧﻭ ﺭﺎﻛﻭﻯﺭﺍﺪﻬﮕﻧﻰﻠﻠﻤﻟﺍﻦﻴﺑﻝﻮﺻﺍ"ﺱﺎﺳﺍﺮﺑﺎﻬﻧﺁﻰﻳﺎﺗ 6-9 ﻯﺎﻫﻩﻭﺮﮔ ﻕﻼﺧﺍﻪﺘﻴﻤﻛﺕﺎﺑﻮﺼﻣﺎﺑﻖﺑﺎﻄﻣﻦﻴﻨﭼﻢﻫﻭ (41) ﻰﻫﺎﮕﺸﻳﺎﻣﺯﺁﻥﺍﻮﻴﺣﺎﺑ
50 ﻢﺠﺣﺭﺩ) ﺪﻴﻤﺳﻼﭘﻡﺮﮔﻭﺮﻜﻴﻣ 100 ﻥﺍﺰﻴﻣﺎﺑ "ﻥﺍﺮﻳﺍﺭﻮﺘﺳﺎﭘﻮﺘﻴﺘﺴﻧﺍ
.ﺪﻧﺪﺷﻦﻤﻳﺍﭗﭼﻯﺎﭘﻒﻛﻪﻴﺣﺎﻧﺭﺩﻯﺪﻠﺟﺮﻳﺯﺕﺭﻮﺻﻪﺑ (ﺮﺘﻴﻟﻭﺮﻜﻴﻣ ﺖﻓﺎﻳﺭﺩﻩﻭﺮﮔﻭﺩﻭﻪﺘﻓﺮﮔﺕﺭﻮﺻﻪﺘﻔﻫ 6 ﻭ 3 ،0 ﻯﺎﻫﻥﺎﻣﺯﺭﺩﺕﺎﻘﻳﺭﺰﺗ ﻥﺍﻮﻨﻋﻪﺑ (pcHﻭpcDNA3.1) پﻮﺗﻰﻠﭘﻥژﺪﻗﺎﻓﻯﺎﻫﺪﻴﻤﺳﻼﭘﻩﺪﻨﻨﻛ .ﺪﻧﺪﺷﻪﺘﻓﺮﮔﺮﻈﻧﺭﺩﻰﻔﻨﻣﻝﺮﺘﻨﻛ
(DTH) ﻯﺮﻴﺧﺄﺗﺖﻴﺳﺎﺴﺣﺩﺎﻳﺩﺯﺍﺦﺳﺎﭘﻰﺳﺭﺮﺑ ﻯﺯﺎﺳﻦﻤﻳﺍﻦﻳﺮﺧﺁﺯﺍﺲﭘﻪﺘﻔﻫ 3 ،(42) ﻰﻠﺒﻗﻩﺪﺷﺵﺭﺍﺰﮔﺵﻭﺭﻖﺒﻃ ﺮﻫﺎﺑﺮﻇﺎﻨﺘﻣپﻮﺗﻰﻠﭘ ﻚﻴﺘﺘﻨﺳﺪﻴﺘﭙﭘﺯﺍﻡﺮﮔﻭﺮﻜﻴﻣ 10 ﻥﺍﺰﻴﻣﺎﻫﺵﻮﻣ
E614+N1406ﺎﻳC132+E405ﻯﺎﻫﺪﻴﺘﭙﭘﻁﻮﻠﺨﻣ،ﺎﻫﻑﺩﺍﺮﺗﺯﺍﻚﻳ
ﻢﺠﺣﺭﺩ،ﻂﺒﺗﺮﻣﺮﻴﻏﺪﻴﺘﭙﭘﻚﻳﺎﻳﻭ (ﺎﻬﻧﺁﺯﺍﻚﻳﺮﻫﻯﻭﺎﺴﻣﻢﺠﺣﻞﻣﺎﺷ) ﻯﺎﻫﻩﻭﺮﮔﺖﺳﺍﺭﻯﺎﭘﻒﻛﻪﺑﻯﺪﻠﺟﺮﻳﺯﺕﺭﻮﺻﻪﺑﺮﺘﻴﻟﻭﺮﻜﻴﻣ 40 ﻰﻠﻛ ﻪﺑﺪﻫﺎﺷﻥﺍﻮﻨﻋﻪﺑﺰﻴﻧﻯژﻮﻟﻮﻳﺰﻴﻓﻡﺮﺳﻯﻭﺎﺴﻣﺭﺍﺪﻘﻣ .ﺪﺷﻖﻳﺭﺰﺗﻩﺪﺷﻦﻤﻳﺍ ﺖﻋﺎﺳ 72 ﻭ 48 ،24 ،18 ﺖﺷﺬﮔﺯﺍﺲﭘﻭﻖﻳﺭﺰﺗﺎﻫﺵﻮﻣﭗﭼﻯﺎﭘﻒﻛ ﻯﺎﻫﻩﻭﺮﮔﺭﺩﺎﻫﺵﻮﻣﭗﭼﻯﺎﭘﺎﺑﻪﺴﻳﺎﻘﻣﺭﺩﺖﺳﺍﺭﻯﺎﭘﻰﮕﺘﺴﺟﺮﺑﺭﺍﺪﻘﻣ .ﺪﺷﻯﺮﻴﮔﻩﺯﺍﺪﻧﺍ (Mitutoyo, Japan) ﺞﻨﺳﺰﻳﺭﻂﺳﻮﺗﻒﻠﺘﺨﻣ
ﺎﻫ
ﻪﺘﻓﺎﻳ
پﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﻤﺳﻼﭘﻰﺣﺍﺮﻃﻭﺎﻫپﻮﺗﻰﭘﺍﺏﺎﺨﺘﻧﺍ
ﺍﺪﺟ ﻯﺎﻫﻪﻧﻮﻤﻧﻪﺑ ﻁﻮﺑﺮﻣCTL ﻯﺎﻫپﻮﺗﻰﭘﺍﻯﺪﻴﺳﺍﻮﻨﻴﻣﺁ ﻰﻟﺍﻮﺗ
ﺖﺳﺍﻩﺪﺷﻱﺯﺍﺪﻧﺍﻩﺍﺭﺭﻮﻈﻨﻣﻦﻳﺍﻱﺍﺮﺑﻪﻛﻰﺗﺎﻋﻼﻃﺍﻩﺎﮕﻳﺎﭘﺭﺩHCVﻩﺪﺷ (http://hcv.lanl.gov/content/immuno/immuno-main.html)
ﻡﺪﻗﺭﺩ .ﺖﺳﺍﻩﺪﺷ ﻯﺯﺍﺪﻧﺍﻩﺍﺭﺭﻮﻈﻨﻣ ﻦﻳﺍﻯﺍﺮﺑﻪﻛ ﺪﺷﺎﺑﻲﻣﺩﻮﺟﻮﻣ ﻰﺣﺍﻮﻧﺯﺍ (43) HLA-A2ﻊﻳﺎﺷﭗﻳﺎﺗﺮﭘﻮﺳﻪﺑ ﻰﮕﺘﺴﺑﺍﻭﺱﺎﺳﺍﺮﺑﻝﻭﺍ
ﻥﺎﻴﻣﺯﺍﻪﻛﺪﻧﺪﺷﻰﻳﺎﺳﺎﻨﺷپﻮﺗﻰﭘﺍ 60 ﺯﺍﺶﻴﺑNS3ﻭCore ،E2
ﻰﻟﺍﻮﺗﺕﺎﺒﺛﺮﻴﻈﻧﻰﻳﺎﻫﻰﮔﮋﻳﻭﺱﺎﺳﺍﺮﺑﺐﻟﺎﻏپﻮﺗﻰﭘﺍ 3 ﺖﻳﺎﻬﻧﺭﺩﺎﻬﻧﺁ ﻡﻭﺎﻘﻣﻭﻊﻳﺎﺷﭗﻴﺗﻮﻧژﻭﺩﺯﺍﻩﺪﺷﺍﺪﺟﻯﺎﻫﻪﻧﻮﻤﻧﻦﻴﺑﺭﺩﻯﺪﻴﺳﺍﻮﻨﻴﻣﺁ ﻯﺯﺎﺴﻛﺎﭘﺎﺑ ﻩﺪﺷ ﺩﺎﺠﻳﺍﻰﻨﻤﻳﺍﺦﺳﺎﭘﻁﺎﺒﺗﺭﺍ ،(9) 1bﻭ 1aﻥﺎﻣﺭﺩﻪﺑ
ﻝﻭﺪﺟ) ﺪﻧﺪﺷ ﺏﺎﺨﺘﻧﺍ HLA-A2.1 ﻪﺑ ﺎﻬﻧﺁﻝﺎﺼﺗﺍ ﻰﺘﻴﻨﻴﻓﺍ ﻭ ﺱﻭﺮﻳﻭ
-140 ﻪﻴﺣﺎﻧ) HLA-A2.1 ﺮﺑﻩﻭﻼﻋC132،ﺎﻫپﻮﺗﻰﭘﺍﻦﻳﺍﻦﻴﺑﺭﺩ .(1
ﻪﺑ .ﺪﺷﻰﻣﻪﺿﺮﻋﻭﻪﺘﺧﺎﻨﺷﺰﻴﻧ (132-142 ﻪﻴﺣﺎﻧ) H-2d ﻂﺳﻮﺗ (132 ﻝﻮﻤﻌﻣﻥﺍﻮﻴﺣﺭﺩﻦﺴﻛﺍﻭﻰﻳﺍﺯﻰﻨﻤﻳﺍﻪﻌﻟﺎﻄﻣﻥﺎﻜﻣﺍﻥﺩﺮﻛﻢﻫﺍﺮﻓﺭﻮﻄﻨﻣ
E405 پﻮﺗ ﻰﭘﺍ ،C132 ﺮﺑ ﻩﻭﻼﻋ (BALB/c ﺵﻮﻣ) ﻰﻫﺎﮕﺸﻳﺎﻣﺯﺁ ﻩﺩﻭﺰﻓﺍ ﻰﺑﺎﺨﺘﻧﺍ ﻯﺎﻫپﻮﺗﻰﭘﺍ ﻉﻮﻤﺠﻣﻪﺑ ﺩﻮﺑH-2d ﻪﺑ ﻪﺘﺴﺑﺍﻭ ﻪﻛ ﺰﻴﻧ ﻰﻨﻤﻳﺍ ﺦﺳﺎﭘ ﺖﺳﺍ ﻩﺪﺷ ﻩﺩﺍﺩ ﻥﺎﺸﻧ 1 ﻝﻭﺪﺟ ﺭﺩ ﻪﻛ ﺭﻮﻃﻥﺎﻤﻫ .ﺪﺷ ﻩﺪﺷ ﺵﺭﺍﺰﮔ ﻒﻠﺘﺨﻣ ﺕﺎﻘﻴﻘﺤﺗ ﺭﺩ ﺎﻫپﻮﺗﻰﭘﺍ ﻦﻳﺍ ﻪﺑ ﺖﺒﺴﻧ ﺐﻟﺎﻏ ﻰﻠﺻﺍﻯﺎﻫپﻮﺗﻰﭘﺍﺯﺍﻰﻜﻳﻥﺍﻮﻨﻋﻪﺑN1406ﻥﺎﻴﻣﻦﻳﺍﺭﺩﻭ (44-49)
ﻪﺑ ﻦﻴﻨﭼﻢﻫ .(49 ،6) ﺖﺳﺍﻩﺪﺷﻪﺘﺧﺎﻨﺷﺱﻭﺮﻳﻭﻯﺯﺎﺳﻙﺎﭘ ﺎﺑﻂﺒﺗﺮﻣ
ﻯﺎﻫﻰﻨﻴﺑﺶﻴﭘﺯﺍﻯﺮﻴﮔﻩﺮﻬﺑﺎﺑ (50) ﺲﻧﺎﻨﻴﻣﻭﺩﻮﻧﻮﻤﻳﺍﻩﺪﻳﺪﭘﺶﻫﺎﻛﺭﻮﻈﻨﻣ
MHC ﻝﻮﻜﻟﻮﻣ ﻪﺑ ﺎﻫپﻮﺗﻰﭘﺍ ﻝﺎﺼﺗﺍ ﻪﺑ ﻁﻮﺑﺮﻣ in silico ﻪﻛ ﺪﺷ ﻰﻌﺳ (http://bio.dfci.harvard.edu/RANKPEP)
-Junc) ﻰﻨﻴﺑﺎﻨﻴﺑﺪﻳﺪﺟ ﻯﺎﻫپﻮﺗﻰﭘﺍ ﺶﻳﺍﺪﻴﭘﻪﻄﺳﺍﻭ ﻪﺑ ﻪﻛﻰﮔﮋﻳﻭ ﺪﻧﺍﻮﺗﻰﻣﻡﻭﺯﺎﺌﺗﻭﺮﭘﻂﺳﻮﺗﺎﻫپﻮﺗﻰﭘﺍﺵﺯﺍﺩﺮﭘﺮﺑﺮﻴﺛﺎﺗﺰﻴﻧﻭ (tional
ﻯﺎﻫﺵﻭﺭﻚﻤﻛﻪﺑ ،(51) ﺩﻮﺷﻊﻗﺍﻭﺮﺛﻮﻣﻰﻨﻤﻳﺍﺦﺳﺎﭘ ﻰﻳﺁﺭﺎﻛﺮﺑ
ﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑ ﺱﺎﺳﺍ ﻦﻳﺍﺮﺑ .ﺖﺳﺍﻰﻨﻴﺑﺶﻴﭘﻞﺑﺎﻗﻚﻴﺗﺎﻣﺭﻮﻔﻧﺍﻮﻧﻮﻤﻳﺍ
ﺶﻴﭘ ﺖﻬﺟ) (52) Rankpep ﻯﺎﻫﻪﻣﺎﻧﺮﺑ ﺭﺩ ﺩﻮﺟﻮﻣﻢﺘﻳﺭﻮﮕﻟﺍ ﻭﺩ
ﺵﺮﺑ ﻰﻨﻴﺑﺶﻴﭘ ﺖﻬﺟ) (53) PAProC ﻭ (MHC ﻯﺎﻫﺪﻧﺎﮕﻴﻟ ﻰﻨﻴﺑ
ﻪﺘﻓﺮﮔﺮﻈﻧﺭﺩپﻮﺗﻰﭘﺍ 4 ﻯﺍﺮﺑﺩﻮﺟﻮﻣﺐﻴﻛﺮﺗ 24 ﻦﻴﺑﺯﺍ (ﻡﻭﺯﺎﺌﺗﻭﺮﭘ ﻭﺮﺘﻤﻛﻰﻨﻴﺑﺎﻨﻴﺑپﻮﺗﻰﭘﺍﻪﻛ (1 ﻞﻜﺷ) ﻑﺩﺍﺮﺗ 3 ،ﻖﻴﻘﺤﺗﻦﻳﺍﺭﺩﻩﺪﺷ ﺭﻮﻛﺬﻣﻯﺎﻫﻰﻟﺍﻮﺗ .ﺪﻧﺪﺷﺏﺎﺨﺘﻧﺍ،ﺪﻧﺩﺍﺩﻰﻣﻥﺎﺸﻧﺍﺭﻯﺮﺘﻬﺑ ﺵﺯﺍﺩﺮﭘ ﻥژﻮﻴﻓ،ﺪﻧﺪﺷﻥﻮﻠﻛpcDNA3.1ﺭﻮﺘﻛﻭﺭﺩﻰﻳﺎﻬﻨﺗﻪﺑﻪﻜﻨﻳﺍﺮﺑﻩﻭﻼﻋ
.ﺪﺷﻥﻮﻠﻛﺪﻴﻤﺳﻼﭘﻦﻳﺍﺭﺩﻪﻧﺎﮔﺍﺪﺟﺭﻮﻃﻪﺑﺰﻴﻧHBsAg-Sﻥژﺎﺑﺎﻬﻧﺁ
ﺖﻳﺎﻬﻧﺭﺩﻭﻰﻤﻳﺰﻧﺁﻯﺎﻫﺰﻴﻟﺎﻧﺁﻂﺳﻮﺗﻩﺪﺷﻪﺘﺧﺎﺳﺪﻴﻤﺳﻼﭘﺶﺷﻪﻴﻠﻛ .ﺪﻨﺘﻓﺮﮔﺭﺍﺮﻗﺪﻴﻳﺎﺗﺩﺭﻮﻣ،ﻰﻟﺍﻮﺗﻦﻴﻴﻌﺗ
پﻮﺗﻰﻠﭘﻯﺎﻫﻥژﻥﺎﻴﺑﻰﺳﺭﺮﺑ
ﻚﻳﺩﻮﺟﻭﻯﺮﺗﻮﻴﭙﻣﺎﻛﻢﺘﻳﺭﻮﮕﻟﺍﻪﻠﻴﺳﻭﻪﺑپﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﺘﭙﭘﺰﻴﻟﺎﻧﺁ ﺭﺩﺺﺧﺎﺷﻭﺩﻭ 3 ﻭ 2 پﻮﺗﻰﻠﭘﺭﺩBﻯﺎﻫﻝﻮﻠﺳﻚﻴﻧژﻰﺘﻧﺁﺺﺧﺎﺷ
،ﻰﻨﻴﺑﺶﻴﭘﻦﻳﺍﺎﺑ ﻮﺳﻢﻫﺰﻴﻧ ﺍﺰﻳﻻﺍﺞﻳﺎﺘﻧ .ﺩﺮﻛﺺﺨﺸﻣﺍﺭ 1 پﻮﺗﻰﻠﭘ ﻡﺮﺳﺭﺩﺍﺭ 1 پﻮﺗﻰﻠﭘﻪﻴﻠﻋﻰﺷﻮﮔﺮﺧﻝﺎﻧﻮﻠﻛﻰﻠﭘﻡﺮﺳﻰﺘﻧﺁﺮﺗﻻﺎﺑﺭﺎﻴﻋ ﺭﺩپﻮﺗﻰﻠﭘ ﻦﻳﺍﺎﺑﺮﻇﺎﻨﺘﻣ ﻚﻴﺘﺘﻨﺳﺪﻴﺘﭙﭘﺎﺑ ﻩﺪﺷ ﻦﻤﻳﺍﻯﺎﻫﺵﻮﮔﺮﺧ ﻩﺪﺸﻧ ﻩﺩﺍﺩﻥﺎﺸﻧﺞﻳﺎﺘﻧ) ﺩﺍﺩ ﻥﺎﺸﻧ 3 ﻭ 2 پﻮﺗﻰﻠﭘ ﻯﺎﻫﺪﻴﺘﭙﭘﺎﺑﻪﺴﻳﺎﻘﻣ
ﻯﺩﺎﺑﻰﺘﻧﺁ ﻭ پﻮﺗﻰﻠﭘ ﺪﻴﺘﭙﭘﻪﺳ ﻪﻴﻠﻋ ﻩﺪﺷ ﻪﻴﻬﺗ ﻯﺎﻫﻡﺮﺳﻰﺘﻧﺁ .(ﺖﺳﺍ ﺭﻮﻈﻨﻣ ﻪﺑ ﻒﻠﺘﺨﻣ ﺕﺎﺸﻳﺎﻣﺯﺁ ﺭﺩ HBsAg ﻪﻴﻠﻋ ﻯﺭﺎﺠﺗ ﻝﺎﻧﻮﻠﻛﻮﻧﻮﻣ
ﻪﻛﺩﺍﺩﻥﺎﺸﻧﺞﻳﺎﺘﻧ .ﺪﻧﺪﺷﻩﺩﺮﺑﺭﺎﻛﻪﺑپﻮﺗﻰﻠﭘﻯﺎﻫﻥژﻝﻮﺼﺤﻣﻰﻳﺎﺳﺎﻨﺷ ﻥژﻰﺘﻧﺁﻦﺘﺷﺍﺩﺎﺑﻪﻛCos-7ﻰﻟﻮﻠﺳﻩﺩﺭﺭﺩﻰﺑﻮﺧﻪﺑپﻮﺗﻰﻠﭘﻯﺎﻫﻥژ
ﺍﺭ pcDNA3.1 ﺪﻴﻤﺳﻼﭘ ﻝﺎﻣﻭﺯﻰﭘﺍ ﺮﻴﺜﻜﺗﻥﺎﻜﻣﺍ SV40ﺱﻭﺮﻳﻭ T
ﻚﻴﻨﻜﺗﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑﻦﻳﺍﺮﺑﻩﻭﻼﻋ .(2 ﻞﻜﺷ) ﺪﻧﺍﻩﺪﺷﻥﺎﻴﺑ،ﺪﻨﻛﻰﻣﻢﻫﺍﺮﻓ ﺪﻳﺮﺒﻴﻫﺐﻴﻛﺮﺗﻮﻧﻯﺎﻫﻦﻴﺌﺗﻭﺮﭘﻥﺎﻴﺑﻪﺑﻁﻮﺑﺮﻣﺪﻧﺎﺑﺕﻼﺑﻥﺮﺘﺳﻭﺩﺭﺍﺪﻧﺎﺘﺳﺍ
.(ﻒﻟﺍ 2 ﻞﻜﺷ) ﺪﺷ ﻩﺪﻫﺎﺸﻣ ﺭﺎﻈﺘﻧﺍ ﺩﺭﻮﻣ ﻩﺯﺍﺪﻧﺍ ﺭﺩ HBsAg-S ﺎﺑ
ﻯﺪﻴﺳﺍﻮﻨﻴﻣﺁﻰﻟﺍﻮﺗﻖﺒﻃHBs-polytopeﺪﻳﺮﺒﻴﻫﻦﻴﺌﺗﻭﺮﭘﻰﻟﻮﻜﻟﻮﻣﻥﺯﻭ
ﺭﺎﻨﻛﺭﺩﺪﻧﺎﺑﻦﻳﺍﺕﻼﺑﻥﺮﺘﺳﻭﺞﻳﺎﺘﻧﺭﺩﻭﺪﺷﺎﺑﻰﻣﻥﻮﺘﻟﺍﺩﻮﻠﻴﻛ 29/8 ﻥﺁ ﻚﻳﺕﺭﻮﺻﻪﺑ ﻥﻮﺘﻟﺍﺩﻮﻠﻴﻛ 33 ﺩﻭﺪﺣﻰﻟﻮﻜﻟﻮﻣﻥﺯﻭ ﺎﺑﻯﺮﮕﻳﺩ ﺪﻧﺎﺑ ﺪﻧﺎﺑﻰﻠﺒﻗﺕﺎﻌﻟﺎﻄﻣﺱﺎﺳﺍﺮﺑﻪﻛﺪﺷﻩﺪﻫﺎﺸﻣ (doublet) ﻪﻧﺎﮔﻭﺩﺪﻧﺎﺑ ﺭﺩ HBsAg ﻦﻴﺌﺗﻭﺮﭘ ﻥﻮﻴﺳﻼﻳﺯﻮﻜﻴﻠﮔ ﻪﺑ ﻁﻮﺑﺮﻣ ﺪﻧﺍﻮﺗﻰﻣ ﺮﺗﻦﻴﮕﻨﺳ
ﺲﻧﺎﺳﺭﻮﻠﻓﻮﻧﻮﻤﻳﺍ .(55 ،54) ﺪﺷﺎﺑﻥﺁ 146 ﻦﻳژﺭﺎﭙﺳﺁﻪﻨﻴﻣﺁﺪﻴﺳﺍﻞﺤﻣ ﻯﺎﻫﻦﻴﺌﺗﻭﺮﭘﻥﺎﻴﺑﺰﻴﻧHBsAgﻪﻴﻠﻋﻯﺩﺎﺑﻰﺘﻧﺁﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑﻢﻴﻘﺘﺴﻣﺮﻴﻏ
ﺕﺎﺒﺛﺍ ﻩﺪﺷ ﺖﻜﻓﺲﻧﺍﺮﺗ Cos-7 ﻯﺎﻫﻝﻮﻠﺳ ﺭﺩ ﺍﺭ HBs-polytope
ﺵﻭﺭﺎﺑﺎﻫﻝﻮﻠﺳﻦﻳﺍﺖﺸﻛﻂﻴﺤﻣﻰﺳﺭﺮﺑﻦﻴﻨﭼﻢﻫ .(ﺏ2 ﻞﻜﺷ) ﺩﺮﻛ ﺩﻮﺑﻝﻮﻠﺳﺖﺸﻛﻂﻴﺤﻣﻪﺑHBsAgﺪﻳﺮﺒﻴﻫﺕﺍﺭﺫﺢﺷﺮﺗﺯﺍﻰﻛﺎﺣﺍﺰﻳﻻﺍ
.(ﺝ2 ﻞﻜﺷ)
ﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑﻰﻟﻮﻠﺳﺕﺍﺰﻴﻟﺭﺩHBs-polytopeﺪﻳﺮﺒﻴﻫﻯﺎﻫﻦﻴﺌﺗﻭﺮﭘﻥﺎﻴﺑﻰﺳﺭﺮﺑ .Cos-7ﻯﺎﻫﻝﻮﻠﺳﺭﺩپﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﺘﭙﭘﺢﺷﺮﺗﻭﻥﺎﻴﺑ :2 ﻞﻜﺷ
ﻩﺪﺷﺕﻼﺑﺎﺸﻏ 2-4 ﻯﺎﻫﻥﻮﺘﺳﺭﺩ .ﺪﺷﻡﺎﺠﻧﺍ (ﺏ،ﻢﻴﻘﺘﺴﻣﺮﻴﻏﺲﻧﺎﺳﺭﻮﻠﻓﻮﻧﻮﻤﻳﺍﻭ (ﻒﻟﺍ،ﺕﻼﺑﻥﺮﺘﺳﻭﺵﻭﺭﻭﺩﺎﺑHBsAgﻪﻴﻠﻋﻝﺎﻧﻮﻠﻛﻮﻧﻮﻣﻯﺩﺎﺑﻰﺘﻧﺁ
ﺪﻨﺷﺎﺑﻰﻣپﻮﺗﻰﻠﭘﺪﻴﻤﺳﻼﭘﻪﺳﺖﻜﻓﺲﻧﺍﺮﺗﺯﺍﻞﺻﺎﺣﻦﻴﺌﺗﻭﺮﭘﻪﻠﻳﺯﻮﻜﻴﻠﮔﺮﻴﻏﻭﻪﻠﻳﺯﻮﻜﻴﻠﮔﻝﺎﻜﺷﺍﻪﻧﺎﺸﻧﻪﻛﻥﻮﺘﻟﺍﺩﻮﻠﻴﻛ 30 ﻭ 33 ﻩﺯﺍﺪﻧﺍﺎﺑﻪﻧﺎﮔﻭﺩﻯﺎﻫﺪﻧﺎﺑ
ﺭﺩ 1 ﻥﻮﺘﺳ) ﺪﻧﺩﺍﺪﻧﻥﺎﺸﻧﺕﻼﺑﻥﺮﺘﺳﻭﺎﺸﻏﺭﺩﺍﺭﻪﻃﻮﺑﺮﻣﺪﻧﺎﺑﺎﻬﻨﺗﻪﻧﻪﺘﻜﻓﺲﻧﺍﺮﺗﺮﻴﻏﻯﺎﻫﻝﻮﻠﺳ .ﺪﻧﺍﻩﺪﺷﺺﺨﺸﻣﻦﻴﻳﺎﭘﻭﻻﺎﺑﺶﻠﻓﻭﺩﺎﺑﺐﻴﺗﺮﺗﻪﺑ
ﻯﺎﻫﻝﻮﻠﺳﺭﺩﺲﻧﺎﺳﺭﻮﻠﻓﺭﻮﻧﻩﺪﻫﺎﺸﻣﻪﻛﺖﺳﺍﻰﻟﺎﺣﺭﺩﻦﻳﺍ .(ﺏﺭﺩb) ﺪﻧﺩﻮﺑﺰﻴﻧﺲﻧﺎﺳﺭﻮﻠﻓﻮﻧﻮﻤﻳﺍﺵﻭﺭﺭﺩﺖﻳﻭﺭﻞﺑﺎﻗﺲﻧﺎﺳﺭﻮﺋﻮﻠﻓﺪﻗﺎﻓﻪﻜﻠﺑ (ﻒﻟﺍ
.(ﺏﺭﺩa) ﺩﻮﺑﺮﻈﻧﺩﺭﻮﻣﻦﻴﺌﺗﻭﺮﭘﻥﺎﻴﺑﻪﻧﺎﺸﻧ (pcHPOL1ﺎﺑﺲﻜﻋﻦﻳﺍﺭﺩ) ﻩﺪﺷﺖﻜﻓﺲﻧﺍﺮﺗ
(ﺝ،ﺪﺷﻰﺳﺭﺮﺑﺎﻫﻝﻮﻠﺳﺖﻜﻓﺲﻧﺍﺮﺗﺯﺍﺪﻌﺑﺖﻋﺎﺳ 72 ﺪﻳﺮﺒﻴﻫﺮﻴﻏHBsAg-SﺎﺑﻪﺴﻳﺎﻘﻣﺭﺩHBs-polytopeﺪﻳﺮﺒﻴﻫﺕﺍﺭﺫﺢﺷﺮﺗﺪﺻﺭﺩﺍﺰﻳﻻﺍﺵﻭﺭﺎﺑ
.ﺪﻨﺷﺎﺑﻰﻣﻒﻠﺘﺨﻣﺖﻜﻓﺲﻧﺍﺮﺗﻪﺳﺯﺍﻞﺻﺎﺣﺭﺎﻴﻌﻣﻑﺍﺮﺤﻧﺍ ±ﻦﻴﮕﻧﺎﻴﻣﺮﮕﻧﺎﻳﺎﻤﻧﺎﻫﻥﻮﺘﺳ
ﺯﺍﻞﺻﺎﺣﮓﻧﺭﺕﺪﺷ .ﺪﻧﺪﺷﻰﺳﺭﺮﺑ (ﺫ،RT-PCRﻭ (ﺩ،ﺕﻼﺑﻮﻧﻮﻤﻳﺍﺕﺍﺩﺵﻭﺭﻭﺩﺎﺑHBsAg-Sﺎﺑﺪﻳﺮﺒﻴﻫﺮﻴﻏپﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﻤﺳﻼﭘﻰﺴﻳﻮﻧﻭﺭﻭﻥﺎﻴﺑ
(3 ﻭ 2 ،1 ﻯﺎﻬﻧﻮﺘﺳﺐﻴﺗﺮﺗﻪﺑ،aﻒﻳﺩﺭ) ﺕﻼﺑﺕﺍﺩﺭﺩ (pcPOL1ﺎﺑﻞﻜﺷﻦﻳﺍﺭﺩ) ﻩﺪﺷﺖﻜﻓﺲﻧﺍﺮﺗﻯﺎﻬﻟﻮﻠﺳﺕﺍﺰﻴﻟﺯﺍﺮﺘﻴﻟﻭﺮﻜﻴﻣ 2ﻭ 10 ،20 ﻯﺭﺍﺬﮔﻪﻧﻮﻤﻧ
ﺪﻴﺘﭙﭘﻡﺮﮔﻮﻧﺎﻧ 4 ﻭ 20 ،100 ﺯﺍﻰﺷﺎﻧﻯﺎﻫﻪﻜﻟﺎﺑﻰﻜﻳﺩﺰﻧﺖﻫﺎﺒﺷﺎﻣﺍﻩﺩﺍﺩﻥﺎﺸﻧ (bﻒﻳﺩﺭ) ﻪﺘﻜﻓﺲﻧﺍﺮﺗﺮﻴﻏﻯﺎﻫﻝﻮﻠﺳﻪﺑﻁﻮﺑﺮﻣﻯﺎﻫﻪﻜﻟﺎﺑﺍﺭﻰﺤﺿﺍﻭﺕﻭﺎﻔﺗ
.ﺖﺷﺍﺩ،ﺪﺷﻪﺘﻓﺮﮔﺮﻈﻧﺭﺩﺖﺒﺜﻣﻝﺮﺘﻨﻛﻥﺍﻮﻨﻋﻪﺑﻪﻛ (3 ﻭ 2 ،1 ﻯﺎﻫﻥﻮﺘﺳﺐﻴﺗﺮﺗﻪﺑ،cﻒﻳﺩﺭ) ﻚﻴﺘﺘﻨﺳ
ﺐﻴﺗﺮﺗﻪﺑ ) 3 ﻭ 2 ،1 پﻮﺗﻰﻠﭘﻯﺎﻫﻥژﺢﻴﺤﺻﻥﺎﻴﺑﻪﺑﻁﻮﺑﺮﻣﻯﺪﻴﺗﻮﺌﻠﻛﻮﻧ 150 ﺪﻧﺎﺑﺭﻮﻀﺣﺰﻴﻧRT-PCRﺭﺩﻩﺪﺷﺖﻜﻓﺲﻧﺍﺮﺗﻯﺎﻫﻝﻮﻠﺳmRNAﺮﻴﺜﻜﺗ
ﻪﻃﻮﺑﺮﻣﺪﻧﺎﺑ (pcDNA3.1) ﻪﻌﻄﻗﻥﻭﺪﺑﺪﻴﻤﺳﻼﭘﺎﺑﻩﺪﺷﺖﻜﻓﺲﻧﺍﺮﺗﻯﺎﻫﻝﻮﻠﺳﻪﺑﻖﻠﻌﺘﻣ (1 ﻥﻮﺘﺳ) ﻝﺮﺘﻨﻛﻪﻧﻮﻤﻧﻪﻛﻰﻟﺎﺣﺭﺩ،ﺩﺍﺩﻥﺎﺸﻧﺍﺭ (2-4 ﻯﺎﻫﻥﻮﺘﺳ
.ﺖﺳﺍﺺﺨﺸﻣﻥﺯﻭﺎﺑﻰﻨﻴﺌﺗﻭﺮﭘﻯﺎﻫﺪﻧﺎﺑﺮﮕﻧﺎﻳﺎﻤﻧMﻥﻮﺘﺳ .ﺖﺷﺍﺪﻧﺍﺭ
ﻒﻟﺍ ﺝ
ﺫ ﺩ
ﺏ
1 2 3 4
kDa 43
34
26
a b
140
120
100
80
60
40
20
0
a
b
c
1 2 3 4 M 1 2 3
Percent
Percent
HBsAg-S
HBs-polytope 1 HBs-polytope 2 HBs-polytope 3
bp 200 150 100
HBs-ﻥﻭﺭﺩﺭﺩﺩﺎﻳﺯﺭﺍﺪﻘﻣﻪﺑHBs-polytopeﺪﻳﺮﺒﻴﻫﺕﺍﺭﺫﺪﻴﻟﻮﺗﻪﭼﺮﮔﺍ
ﺲﻧﺎﺳﺭﻮﻠﻓﻮﻧﻮﻤﻳﺍ ﻭﺕﻼﺑﻥﺮﺘﺳﻭﻯﺎﻫﺵﻭﺭﺎﺑﻭﻪﺘﻓﺮﮔﺕﺭﻮﺻﻝﻮﻠﺳ ﻂﺳﻮﺗﻩﺪﺷ ﺪﻴﻟﻮﺗ) ﻰﻌﻴﺒﻃHBsAgﺕﺍﺭﺫﺎﺑﻪﺴﻳﺎﻘﻣ ﺭﺩﺎﻣﺍﺪﺷﺪﻴﻳﺎﺗ
ﺩﻮﺑﺮﺘﻤﻛﺖﺸﻛﻂﻴﺤﻣﻪﺑﺪﻳﺮﺒﻴﻫﺕﺍﺭﺫﺢﺷﺮﺗﻥﺍﺰﻴﻣ (pcHﺪﻴﻤﺳﻼﭘ
.(ﺪﺻﺭﺩ 30 ﺩﻭﺪﺣ)
ﻩﺯﺍﺪﻧﺍ HBsAg ﺎﺑ ﺪﻳﺮﺒﻴﻫﺮﻴﻏ پﻮﺗﻰﻠﭘ ﻯﺎﻫﺪﻴﺘﭙﭘ ﻪﻛ ﺎﺠﻧﺁ ﺯﺍ
(ﻥﻮﺘﻟﺍﺩﻮﻠﻴﻛ 4 ﺩﻭﺪﺣﻰﻟﻮﻜﻟﻮﻣﻥﺯﻭﺎﺑﻪﻨﻴﻣﺁﺪﻴﺳﺍ 40) ﻪﺘﺷﺍﺩﻚﭼﻮﻛ ﻥﺎﻴﺑ ،ﺩﻮﺒﻧ ﺭﺎﻴﺘﺧﺍ ﺭﺩ ﺰﻴﻧ ﺎﻬﻧﺁﻪﻴﻠﻋ ﻯﺭﺎﺠﺗ ﻝﺎﻧﻮﻠﻛﻮﻧﻮﻣﻯﺩﺎﺑﻰﺘﻧﺁ ﻭ
RT-PCR ﻭ (ﺩ2 ﻞﻜﺷ) ﺕﻼﺑﺕﺍﺩﻯﺎﻫﻚﻴﻨﻜﺗ ﺯﺍ ﻩﺩﺎﻔﺘﺳﺍ ﺎﺑ ﺎﻬﻧﺁ ﻥﺎﺸﻧ mRNA ﻰﺴﻳﻮﻧﻭﺭ ﻭ ﻪﻤﺟﺮﺗ ﺢﻄﺳ ﺭﺩ ﺐﻴﺗﺮﺗ ﻪﺑ (ﺫ2 ﻞﻜﺷ) ﻪﺑ ﻁﻮﺑﺮﻣ ﻯﺎﻫﻪﻜﻟ ﮓﻧﺭ ﺕﺪﺷ ﺖﻫﺎﺒﺷ ﺕﻼﺑ ﺕﺍﺩ ﺭﺩ .ﺪﺷ ﻩﺩﺍﺩ ﻯﺎﻫﺪﻴﺘﭙﭘﺯﺍﻩﺪﺷﻯﺭﺍﺬﮔﻪﻧﻮﻤﻧﻯﺎﻫﻪﻜﻟﺎﺑﻩﺪﺷﺖﻜﻓﺲﻧﺍﺮﺗﻯﺎﻫﻝﻮﻠﺳ ﻯﺎﻫﻝﻮﻠﺳﻯﺎﻫﻪﻜﻟﺎﺑﺎﻬﻧﺁﺺﺨﺸﻣﺕﻭﺎﻔﺗﻭ (ﺖﺒﺜﻣﻝﺮﺘﻨﻛ) ﻚﻴﺘﺘﻨﺳ ﺭﺩپﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﻤﺳﻼﭘﻥﺎﻴﺑﺯﺍﻰﻛﺎﺣ (ﻰﻔﻨﻣﻝﺮﺘﻨﻛ) ﻪﺘﻜﻓﺲﻧﺍﺮﺗﺮﻴﻏ ﻡﺎﺠﻧﺍﺯﺍﻰﺷﺎﻧﻱﺪﻴﺗﻮﺌﻠﻛﻮﻧ 150 ﺪﻧﺎﺑﺩﻮﺟﻭﻦﻴﻨﭼﻢﻫ .ﺩﻮﺑﺎﻫﻝﻮﻠﺳﻦﻳﺍ ﻯﺎﻫﺪﻴﻤﺳﻼﭘﺎﺑﻩﺪﺷﺖﻜﻓﺲﻧﺍﺮﺗﻯﺎﻫﻝﻮﻠﺳRNAﻯﻭﺭﺮﺑRT-PCR
ﻩﺪﺷﺖﻜﻓﺲﻧﺍﺮﺗﻯﺎﻫﻝﻮﻠﺳﻪﻧﻮﻤﻧﺭﺩﺪﻧﺎﺑﻦﻳﺍﻩﺪﻫﺎﺸﻣﻡﺪﻋﻭپﻮﺗﻰﻠﭘ ﻞﻜﺷﺭﺩ) RTﻢﻳﺰﻧﺁﺪﻗﺎﻓ ﻝﺮﺘﻨﻛﻪﻧﻮﻤﻧﺰﻴﻧﻭﻪﻌﻄﻗﻥﻭﺪﺑﺪﻴﻤﺳﻼﭘﺎﺑ
پﻮﺗﻰﻠﭘ ﻯﺎﻫﻥژ ﺯﺍﻰﺴﻳﻮﻧﻭﺭ ﻩﺪﻨﻨﻛ ﺪﻴﻳﺎﺗ ،(ﺖﺳﺍ ﻩﺪﺸﻧ ﻩﺩﺍﺩ ﻥﺎﺸﻧ
.ﺩﻮﺑﺮﻈﻧﺩﺭﻮﻣ
BALB/cﺵﻮﻣﺭﺩDTHﺦﺳﺎﭘ
CTL ﻯﺎﻫپﻮﺗﻰﭘﺍ ﻪﻴﻠﻋ ﻰﺻﺎﺼﺘﺧﺍ ﻰﻨﻤﻳﺍ ﺦﺳﺎﭘ ﻪﻴﻟﻭﺍ ﻰﺑﺎﻳﺯﺭﺍ DTHﻰﺘﺳﻮﭘﻥﻮﻣﺯﺁﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑپﻮﺗﻰﻠﭘﻯﺎﻫﻩﺯﺎﺳﺭﺩﻩﺪﺷﻩﺪﻧﺎﺠﻨﮔ
(42) ﺖﺳﺍin vivoﺭﺩﻰﻟﻮﻠﺳﻰﻨﻤﻳﺍﻰﺳﺭﺮﺑﺩﺭﺍﺪﻧﺎﺘﺳﺍﺵﻭﺭﻚﻳﻪﻛ
-ﻚﻴﺘﺘﻨﺳﻯﺎﻫﺪﻴﺘﭙﭘ ﻖﻳﺭﺰﺗﺯﺍﺲﭘﺭﻮﻈﻨﻣﻦﻳﺍﻯﺍﺮﺑ .ﺖﻓﺮﮔﺕﺭﻮﺻ -ﻭﺩﺎﺑﺮﻇﺎﻨﺘﻣﻯﺎﻫﺪﻴﺘﭙﭘﻁﻮﻠﺨﻣ،ﻒﻠﺘﺨﻣﻯﺎﻫﻑﺩﺍﺮﺗﺎﺑﺮﻃﺎﻨﺘﻣپﻮﺗﻰﻠﭘ ﻰﻧﺎﺴﻧﺍپﻮﺗﻰﭘﺍﻭﺩﺎﺑﺮﻇﺎﻨﺘﻣﻯﺎﻫﺪﻴﺘﭙﭘﻁﻮﻠﺨﻣﺎﻳ (H-2d) ﻰﺷﻮﻣپﻮﺗﻰﭘﺍ ﻡﺭﻮﺗﻥﺍﺰﻴﻣﺖﻋﺎﺳ 24 ﺯﺍﺲﭘ،ﺎﻫﺵﻮﻣﺖﺳﺍﺭﻯﺎﭘﻒﻛﻪﺑ (HLA-A2) ﻩﺪﺷﻖﻳﺭﺰﺗﻯژﻮﻟﻮﻳﺰﻴﻓﻡﺮﺳﻯﻭﺎﺴﻣﻢﺠﺣﺎﺑﻪﻛﭗﭼﻯﺎﭘﺎﺑﻪﺴﻳﺎﻘﻣﺭﺩﺎﭘ
.(3 ﻞﻜﺷ) ﺪﺷﻯﺮﻴﮔﻩﺯﺍﺪﻧﺍﺩﻮﺑ
پﻮﺗﻰﻠﭘﺪﻴﺘﭙﭘﺎﺑﻩﺪﺷﻚﻳﺮﺤﺗﻯﺎﻫﺵﻮﻣﻦﻴﺑﻪﻨﻴﺴﻛﺍﻭﻩﻭﺮﮔﺮﻫﺭﺩ ﻚﻳﺮﺤﺗﻯﺎﻫﺵﻮﻣﻭ (E405+C132) ﻰﺷﻮﻣپﻮﺗﻰﭘﺍﻭﺩﻁﻮﻠﺨﻣﺎﻳ ﺪﺷ ﻩﺪﻫﺎﺸﻣ ﻯﺭﺍﺩﻰﻨﻌﻣ ﺕﻭﺎﻔﺗ (ﻂﺒﺗﺮﻣﺮﻴﻏ) ﻝﺮﺘﻨﻛ ﺪﻴﺘﭙﭘ ﺎﺑ ﻩﺪﺷ ﻪﻴﻠﻋ ﻩﺪﺷ ﺩﺎﺠﻳﺍ ﻰﻟﻮﻠﺳ ﺦﺳﺎﭘﻥﺩﻮﺑ ﻰﺻﺎﺼﺘﺧﺍ ﻪﻧﺎﺸﻧﻪﻛ (p<0/01) ﺩﻮﺑﻰﻟﺎﺣﺭﺩﻦﻳﺍ .ﺩﻮﺑﻰﺷﻮﻣﻯﺎﻫپﻮﺗﻰﭘﺍﻁﻮﻠﺨﻣﺰﻴﻧﻭپﻮﺗﻰﻠﭘﺪﻴﺘﭙﭘ ﻁﻮﻠﺨﻣﺎﺑﻩﺪﺷﻚﻳﺮﺤﺗﻯﺎﻫﺵﻮﻣﺭﺩﺭﺍﺩﻰﻨﻌﻣﻰﮕﺘﺴﺟﺮﺑﺶﻳﺍﺰﻓﺍﻪﻛ ﺭﺩ ﻦﻴﻨﭽﻤﻫ .ﺪﺸﻧ ﻩﺪﻫﺎﺸﻣ (E614+N1406) ﻰﻧﺎﺴﻧﺍ ﻯﺎﻫپﻮﺗﻰﭘﺍ
pcDNA3.1، pcH ﺪﻴﻤﺳﻼﭘﺎﺑﻪﻨﻴﺴﻛﺍﻭ) ﻝﺮﺘﻨﻛﻯﺎﻫﻩﻭﺮﮔﺎﺑﻪﺴﻳﺎﻘﻣ ﻥﺎﻣﺭﺩﻯﺎﻫﻩﻭﺮﮔ (ﺮﺘﻣﻰﻠﻴﻣ 0–0/03 ﻡﺭﻮﺗﻦﻴﮕﻧﺎﻴﻣﺎﺑ،ﻪﻨﻴﺴﻛﺍﻭﺮﻴﻏﺎﻳﻭ ﺎﺑ) ﻰﺷﻮﻣپﻮﺗﻰﭘﺍﻭﺩﻁﻮﻠﺨﻣﺎﻳپﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﺘﭙﭘﺎﺑﻩﺪﺷﻚﻳﺮﺤﺗ ﺶﻳﺍﺰﻓﺍ (ﺮﺘﻣﻰﻠﻴﻣ 0/11-0/2 ﻭ 0/2-0/46 ﺐﻴﺗﺮﺗ ﻪﺑ ﻡﺭﻮﺗ ﻦﻴﮕﻧﺎﻴﻣ ﻪﻜﻨﻳﺍﻪﺟﻮﺗﺐﻟﺎﺟﻪﺘﻜﻧ .(p<0/01) ﺪﻧﺩﺍﺩﻥﺎﺸﻧﺍﺭﻯﺭﺍﺩﻰﻨﻌﻣﻰﮕﺘﺴﺟﺮﺑ ﻯﺎﻫﺪﻴﻤﺳﻼﭘﺎﺑﻩﺪﺷﻦﻤﻳﺍﻯﺎﻫﻩﻭﺮﮔﺭﺩﺎﭘﻰﮕﺘﺴﺟﺮﺑﺶﻳﺍﺰﻓﺍﻦﻴﮕﻧﺎﻴﻣ
ﻭ (ﺮﺘﻣﻲﻠﻴﻣ 0/36) pcHPOL2 ،(ﺮﺘﻣﻲﻠﻴﻣ 0/33) pcHPOL1
ﻪﻨﻴﺴﻛﺍﻭ ﻯﺎﻫﻩﻭﺮﮔ ﺎﺑ ﻪﺴﻳﺎﻘﻣ ﺭﺩ (ﺮﺘﻣﻲﻠﻴﻣ 0/46) pcHPOL3
ﺪﻧﺩﻮﺑ HBsAg-S ﻥژ ﺪﻗﺎﻓ ﻪﻛ ﻪﻃﻮﺑﺮﻣ پﻮﺗﻰﻠﭘ ﻯﺎﻫﺪﻴﻤﺳﻼﭘ ﺎﺑ
0/3 ﻭ 0/24 ،0/2 ﺎﺑﺐﻴﺗﺮﺗﻪﺑ،pcPOL3ﻭpcPOL1، pcPOL2)
.(p<0/05) ﺖﺷﺍﺩﻯﺭﺍﺩﻰﻨﻌﻣﺕﻭﺎﻔﺗﺰﻴﻧ (ﺮﺘﻣﻰﻠﻴﻣ
.پﻮﺗﻰﻠﭘ ﻲﻧژﻯﺎﻫﻩﺯﺎﺳﺎﺑﻩﺪﺷﻦﻤﻳﺍﻯﺎﻫﺵﻮﻣDTHﺦﺳﺎﭘ :3 ﻞﻜﺷ
ﻰﻟﺍﻮﺗﻒﻠﺘﺨﻣﻯﺎﻫﻑﺩﺍﺮﺗﻩﺪﻨﻨﻛﺪﻛﻯﺎﻫﺪﻴﻤﺳﻼﭘﺎﺑBALB/cﻯﺎﻫﺵﻮﻣ
ﺯﺍﺪﻌﺑﻪﺘﻔﻫﻪﺳ .ﺪﻧﺪﺷﻦﻤﻳﺍHBsAg-Sﻪﺑﻞﺼﺘﻣﺮﻴﻏﺎﻳﻞﺼﺘﻣپﻮﺗﻰﻠﭘ
ﺎﻳﻭﻰﻋﺎﻨﺻپﻮﺗﻰﻠﭘﺪﻴﺘﭙﭘﻡﺮﮔﻭﺮﻜﻴﻣ 10 ﻂﺳﻮﺗﺎﻫﺵﻮﻣﻖﻳﺭﺰﺗﻦﻳﺮﺧﺁ
E614 + N1406ﺎﻳC132 + E405ﻰﭘﻮﺗﻰﭘﺍﻯﺎﻫﺪﻴﺘﭙﭘﺯﺍﻯﻭﺎﺴﻣﻁﻮﻠﺨﻣ
ﺯﺍﺲﭘﻭﻩﺪﺷﻚﻳﺮﺤﺗﻯﺪﻠﺟﺮﻳﺯﺕﺭﻮﺻﻪﺑﺖﺳﺍﺭﻯﺎﭘﻒﻛﻪﻴﺣﺎﻧﺭﺩ
ﻪﻠﻴﺳﻮﺑﻩﺪﺷﻖﻳﺭﺰﺗﭗﭼﻯﺎﭘﺎﺑﻪﺴﻳﺎﻘﻣﺭﺩﻩﺪﺷﺩﺎﺠﻳﺍﻡﺭﻮﺗﺖﻋﺎﺳ 24
ﻯﺎﻫﻩﻭﺮﮔﺰﻴﻧﻭﻦﻤﻳﺍﺮﻴﻏﻯﺎﻫﺵﻮﻣ .ﺪﺷﻯﺮﻴﮔﻩﺯﺍﺪﻧﺍﻯژﻮﻟﻮﻳﺰﻴﻓﻡﺮﺳ
ﻥﺍﻮﻨﻋ ﻪﺑ(pcH ﻭ pcDNA3.1)ﻪﻌﻄﻗ ﺪﻗﺎﻓﻯﺎﻫﺪﻴﻤﺳﻼﭘ ﺎﺑ ﻪﻨﻴﺴﻛﺍﻭ
ﻪﺑﻖﻠﻌﺘﻣ) ILKEPVHGV ﺪﻴﺘﭙﭘﺯﺍ .ﺪﻧﺪﺷﻪﺘﻓﺮﮔﺮﻈﻧﺭﺩﻰﻔﻨﻣﻝﺮﺘﻨﻛ
ﺞﻳﺎﺘﻧ .ﺪﺷﻩﺩﺎﻔﺘﺳﺍﻂﺒﺗﺮﻣﺮﻴﻏﺪﻴﺘﭙﭘﻥﺍﻮﻨﻋﻪﺑ (HIVﺱﻭﺮﻳﻭRTﻢﻳﺰﻧﺁ
(ﺩﺪﻋ 6-9) ﻩﻭﺮﮔﺮﻫﻯﺎﻫﺵﻮﻣﺭﺩﻩﺪﺷﺩﺎﺠﻳﺍﻡﺭﻮﺗﻦﻴﮕﻧﺎﻴﻣﻩﺪﻨﻫﺩﻥﺎﺸﻧ
ﺖﻓﺎﻳﺭﺩﻯﺎﻫﻩﻭﺮﮔﻯﺎﭘﻡﺭﻮﺗﺭﺍﺩﻰﻨﻌﻣﺕﻭﺎﻔﺗ * ﻭﻩﺩﻮﺑﺩﺭﺍﺪﻧﺎﺘﺳﺍﻯﺎﻄﺧ ±
ﺎﺑ ﻪﻨﻴﺴﻛﺍﻭﻯﺎﻫﻩﻭﺮﮔﺎﺑ ﻪﺴﻳﺎﻘﻣﺭﺩﺍﺭpcHPOLﻯﺎﻫﺪﻴﻤﺳﻼﭘﻩﺪﻨﻨﻛ
.ﺪﻫﺩﻰﻣﻥﺎﺸﻧpcPOLﻥژﻮﻴﻓﺮﻴﻏﻯﺎﻫﺪﻴﻤﺳﻼﭘ
ﺚﺤﺑ
ﺯﺍﻯﺪﻳﺪﺟﻞﺴﻧﻥﺍﻮﻨﻋﻪﺑﻥﺍﻮﺗﻰﻣﺍﺭپﻮﺗﻰﻠﭘDNAﻯﺎﻫﻦﺴﻛﺍﻭ
.ﺖﻓﺮﮔ ﺮﻈﻧﺭﺩ ،ﺪﻨﺷﺎﺑﻰﻣ ﻯﺩﺎﻳﺯ ﻩﻮﻘﻟﺎﺑ ﻯﺎﻳﺍﺰﻣﻯﺍﺭﺍﺩ ﻪﻛ ﺎﻫﻦﺴﻛﺍﻭ ﻚﻳﻯﺎﻫپﻮﺗﻰﭘﺍﻪﺿﺮﻋﻭﺵﺯﺍﺩﺮﭘ،ﺏﺬﺟﻥﺎﻳﺮﺟﺭﺩﻪﻛﺎﺠﻧﺁﺯﺍﺎﻣﺍ ﻝﻮﻠﺳﻥﻭﺭﺩﺭﺩﻡﻭﺯﺎﺌﺗﻭﺮﭘﺵﺮﺑﻭﺮﺛﺍﻩﻮﺤﻧﻪﺑﻁﻮﺑﺮﻣﺕﺎﻴﺋﺰﺟ،ﻥژﻰﺘﻧﺁ ﻼﻣﺎﻛﺯﻮﻨﻫپﻮﺗﻰﭘﺍﻚﻳﺵﺯﺍﺩﺮﭘﺮﺑﺭﻭﺎﺠﻣﻪﻨﻴﻣﺁﻯﺎﻫﺪﻴﺳﺍﺮﻴﺛﺎﺗﺰﻴﻧﻭ ﺖﻠﻋﻪﺑپﻮﺗﻰﻠﭘﻯﺎﻫﺪﻴﺘﭙﭘﻥﺎﻴﺑﺮﮕﻳﺩﻑﺮﻃﺯﺍﻭ،(22) ﺖﺴﻴﻧﺺﺨﺸﻣ ،23) ﺪﺷﺎﺑﻰﻣﻒﻴﻌﺿﻡﻮﺳﻭﻡﻭﺩﻯﺎﻫﻥﺎﻤﺘﺧﺎﺳﻥﺍﺪﻘﻓﻭﻥﺩﻮﺑﻚﻴﺘﺘﻨﺳ ﻭﻥﺎﻴﺑﺶﻳﺍﺰﻓﺍﺖﻬﺟﻰﺗﺍﺪﻴﻬﻤﺗﻯﺮﻴﮔﺭﺎﻛﻪﺑﻭﺐﺳﺎﻨﻣﻰﺣﺍﺮﻃ،(28 ،27 ﺩﺭﺍﻮﻣﻪﺑﻪﺟﻮﺗﺎﺑ .ﺖﺳﺍﺖﻴﻤﻫﺍﺰﻳﺎﺣﺭﺎﻴﺴﺑﻦﺴﻛﺍﻭﻉﻮﻧﻦﻳﺍﻰﻳﺍﺯﻰﻨﻤﻳﺍ ﻪﻴﻠﻋپﻮﺗﻰﻠﭘﻯﺎﻫﻩﺯﺎﺳﻰﺣﺍﺮﻃﺭﺩﻪﻛﺩﺍﺩﻥﺎﺸﻧﻖﻴﻘﺤﺗﻦﻳﺍﺞﻳﺎﺘﻧﻕﻮﻓ ﻰﻳﺎﻫﺭﺎﻜﻫﺍﺭ ﺯﺍﻩﺩﺎﻔﺘﺳﺍﺰﻴﻧﻭ ﻚﻴﺗﺎﻣﺭﻮﻔﻧﺍﻮﻧﻮﻤﻳﺍﻯﺎﻫﻰﻨﻴﺑﺶﻴﭘ،HCV
ﻭﺐﺳﺎﻨﻣﻥﺎﻴﺑﻪﺑﺪﻧﺍﻮﺗﻰﻣﺎﻫﻩﺯﺎﺳﻲﻳﺍﺯﻰﻨﻤﻳﺍﻭﻥﺎﻴﺑﻯﺯﺎﺳﻪﻨﻴﻬﺑﺖﻬﺟ .ﺩﻮﺷﺮﺠﻨﻣﻦﺴﻛﺍﻭﺭﺩﻩﺪﺷﻩﺪﻧﺎﺠﻨﮔﻯﺎﻫپﻮﺗﻰﭘﺍﻰﻳﺍﺯﻰﻨﻤﻳﺍﺶﻳﺍﺰﻓﺍ ،پﻮﺗﻰﻠﭘﻯﺎﻫﻩﺯﺎﺳﻰﺣﺍﺮﻃﺭﺩﻩﺪﺷﻪﺘﻓﺮﮔﺮﻈﻧﺭﺩﺕﺎﻜﻧﻪﻠﻤﺟﺯﺍ ﻯﺎﻫﻝﻮﻠﺳﺭﺩﻪﻤﺟﺮﺗﺶﻳﺍﺰﻓﺍﺭﻮﻈﻨﻣﻪﺑ ﻰﻜﻴﺘﻧژﻯﺎﻫﻥﻭﺪﻛﻯﺯﺎﺳﻪﻨﻴﻬﺑ ﺮﺛﻮﻣDNAﻯﺎﻫﻦﺴﻛﺍﻭﻰﻳﺍﺯﻰﻨﻤﻳﺍﺶﻳﺍﺰﻓﺍﺭﺩﺪﻧﺍﻮﺗﻰﻣﻪﻛﺩﻮﺑﻯﺭﻮﻧﺎﺟ
ﻰﻟﺍﻮﺗﻪﺳﺎﺑﺮﻇﺎﻨﺘﻣﻯﺪﻴﺗﻮﺌﻠﻛﻮﻧﺲﻧﺎﻜﺳﺭﻮﻈﻨﻣﻦﻳﺍﻯﺍﺮﺑ .(56) ﺪﺷﺎﺑ ﻪﻣﺎﻧﺮﺑﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑﻩﺪﺷﻰﺣﺍﺮﻃپﻮﺗﻰﻠﭘ
Sequence Manipulation Suite (http://bio.dfci.harvard. edu/Tools/SMS/rev_trans.html)
ﻰﻧﺎﺴﻧﺍﻯﺎﻫﻥﻭﺪﻛﻰﻧﺍﻭﺍﺮﻓﻝﻭﺪﺟﺱﺎﺳﺍﺮﺑﻭ
(www.kazusa.org.jp/codon/)
ﻯﺎﻫﻝﻮﻠﺳﺭﺩﺞﻳﺍﺭﻯﺎﻫﻥﻭﺪﻛﻪﺑﻰﻜﻴﺘﻧژﻯﺎﻫﻥﻭﺪﻛﺮﻴﻴﻐﺗ .ﺪﺷﻦﻴﻴﻌﺗ ﻥﺍﻮﻨﻋﻪﺑﺍﺪﺘﺑﺍﻪﻠﺻﺎﺣﻪﻨﻴﻣﺁﻯﺎﻫﺪﻴﺳﺍﻰﻟﺍﻮﺗﺭﺩﺮﻴﻴﻐﺗﺩﺎﺠﻳﺍﻥﻭﺪﺑﻯﺭﻮﻧﺎﺟ ﻰﺋﺎﻳﺮﺘﻛﺎﺑ ﻯﺎﻫﻥژ ﻯﻭﺎﺣ ﻯﺎﻫﻩﺯﺎﺳ ﺭﺩ ﻥﺎﻴﺑﺶﻳﺍﺰﻓﺍ ﻯﺎﻫﻩﺍﺭ ﺯﺍ ﻰﻜﻳ ﻦﻴﺷﺎﻣﺯﺍﻰﻌﻴﺒﻃﺭﻮﻃﻪﺑﻰﺳﻭﺮﻳﻭﻯﺎﻫﻥژﻪﻜﻨﻳﺍﻪﺑﻪﺟﻮﺗﺎﺑﻭﺩﻮﺑﺡﺮﻄﻣ
60
50
40
30
20
10
0
Control peptide Polytope peptide
E405+C132 E614+N1406
pcPOL1 pcDNA
pcH pcPOL2pcPOL3pcHPOL1
Footpad Swelling (x10
-2 mm)
pcHPOL2pcHPOL3
