B 11 ..

... .. . .. .. , - 1543 2011 .

() , .

2 ( 200 ).

, .

B .

, , , .

: , , , .

.

, , . .

- .

, .

,

(, , Ig) , , , . IgG 70% . IgA 20%. IgM - 10%. . .CDR , CDR . CDR3 . 108 .

, V- J- D .

CDR3 V- (123-129 ), D (27) J (9) .

V D J .

B , , . .. .

1. -.

2. , , ( 8-10 ) (Invitrogen). .

3. -- .

4.

3- 5. 3' , - . - , . IgE IgD , - IgA, IgG, IgM

- , in vitro () ().

:1. .2. .3. . 4. 5. .

:1. 9496 C(5 20). 2. . (30-2)3. . , .

5.Nested () Nested . , . . 6 V-, . 18 (36V) . J- , , . (, AAAA IgA, )

. . , , . , , . , .

- ( ). 1- ; 2 ; 3 ; 4 ; 5 .

. , , . , , . , , - , . . , , . . Roche (454) Genome Sequencer FLX

. .

HCG-rt GTGTTGCTGGGCTTGTGHCM-rt CTGTGAGGTGGCTGCGTIGA-rt GCTGGCTGCTCGTGGTGT

HV1CTGGRGCTGAGGTGAAGAAGHV2CAGRTCACCTTGARGGAGTCTGHV3CGAGGTGCAGCTGKTGGAGWCCHV4TCRSAGACCCTGTCCCTCACCTGCHV5GCAGTCTGGAGCAGAGGTGAHV6TCTCACTCACCTGTGCCATCTCCG

HCG-nested1GAAGTAGTCCTTGACCAGGCAHCM-nested1GTGATGGAGTCGGGAAGGAAGA-nested1 GCGACGACCACGTTCCCATCT

IGHV3GAARGGCMGRTTCACCATCTCIGHV3 GTATCTKCAAATGAACAGYCTGAIGHV1CACAGCCTACATGGAGCTGAGIGHV2GACCAACATGGACCCTGTGGAIGHV4 CCAGTTCTCCCTGAAGCTGAGIGHV5 GTCCATCAGCACYGCCTACCTIGHV6CCAGACACATCCAAGAACCAG

IGHJ-1aAAAAGAGGAGACGGTGACCRKGGTIGHJ-1g GAAAGAGGAGACGGTGACCRKGGTIGHJ-1m CAAAGAGGAGACGGTGACCRKGGTIGHJ-2a AGAAGAGGAGACGGTGACCRKGGTIGHJ-2g GGAAGAGGAGACGGTGACCRKGGTIGHJ-2m CGAAGAGGAGACGGTGACCRKGGTIGHJ-3a ACAAGAGGAGACGGTGACCRKGGTIGHJ-3g GCAAGAGGAGACGGTGACCRKGGTIGHJ-3m CAAGAGGAGACGGTGACCRKGGT

G, 2 G, 1 G, 3

, 1 , 2 , 3

, 1 , 2 , 3

, V-.

, , , , .

: , , , , , . .

.

.

, . .. , .. .

1) .. " ". 1991. 6 . ( 1988.) .."" , 1998.2) ., ., . , , 1990.3) " " . . , . . , , 2002. 4) ( . ..). , . .., 1998.5) ( .). , ,1997.6) . . N 1996, 1.-.60-61.7) Teksam M, Tali T, Kocer B, Isik S. Qualitative and quantitative volumetric evaluation of the efficacy of intravenous immunoglobulin in multiple sclerosis: preliminary report. Neuroradiology 2000, 42(12)8) .., .. . : ( . .., ..) 2000.9) .., .., .. . , - 1997.10) .. . , , 2003.11) Arnason B.et al. Neutralizing antibodies: are they an issue? International MS Journal. 1997.12) K.Murphy, P.Travers, M.Walport Janeway's Immunobiology, 2007. Edition 7th. 13) . . , , 1998.14) Matz M, Shagin D, Bogdanova E, Britanova O, Lukyanov S, Diatchenko L, Chenchik A (1999) Amplification of cDNA ends based on template-switching effect and step-out PCR. Nucleic Acids Res. 27, 1558-1560.15) Strasser-Fuchs S, Fazekas F, Deisenhammer F et al. The Austrian Immunoglobulin in MS (AIMS) study: final analysis. Mult Scler 2000.-:16) International ImMunoGeneTics information system (http://www.imgt.org).17) United States National Library of Medicine. NCBI (National Center for Biotechnology Information). (http://www.ncbi.nlm.nih.gov/pubmed)18) Basic Local Alignment Search Tool. NCBI (National Center for Biotechnology Information). (http://blast.ncbi.nlm.nih.gov/Blast.cgi)

1. 100 , , ( 8-10 ) 500 (Invitrogen).

.2. 200 .3. 10 . 12000 /., . , , . . () , (). 4. . 600 (100%) 1 ( , ). 5. 10 . 12000 /, . , . 6. . 1000 (80%).7. 5 . 7500 /. .8. ( 10 ).9. 10 .

1. , : ( 0.25 2 ) 3 3` (HCArt1/HCGrt1/HCMrt1) (10 ) 1 MQ 1 5 .2. (Microspin FV 2400, BIOSAN).3. ( , ) 2 70 .4. 42 2 .5. , : 5x 2 Mint () 1 dNTP (10 mM) 1 DTT ( 20 mM) 1 5 .6. .7. 5 . 10 .8. .9. 20 , . 10. 42 1 11. 4, . .

MJ Mini BIORAD. : MQ 19,5 10 buffer for Encyclo pol. 2,5 1 50 Encyclo pol. 0,5 V (10 ) 0,5 (10 ) 0,5 50 dNTP 0,5 25 . , 1 95C. 18 :1. , 10 . 95C.2. , 20 . 62C.3. , 10 . 72C., , 4C.

() MQ 20 , . : MQ 19,5 10 buffer for Encyclo pol. 2,5 - 1 1 50 Encyclo pol. 0,5 V (10 ) 0,5 J (10 ) 0,5 50 dNTP 0,5 25 , , , - 64C. 28 25 .

1,2%

1. 0,36 2. 30 1 -- (), 0,5 /.3. 4. - . , 1,5 .

( 10-15 ) . - (. .10). 1 0,5 /, , . , ( Orange G 2%, 30%).

54 (3 3 6 V-) 6 , V . , 25 , 225 .

, :1. 22,5 3 , 675 96% 0,5 .2. . 3. 10 13400 /.4. . ( ), .5. 800 80% . .6. 5 13400 /. 7. . 8. 15 MQ. 10 . 65C .

QIA quick Gel Extraction Kit (Qiagen).

1. Buffer Q6. 3 1 (100~100).2. 10 . 50C, .3. 1 (100%).4. , 1 13400 /. , . 5. 0,5 Buffer Q6.6. 1 13400 /. , . 7. 0,75 Buffer PE, 3 .8. 1 13400 /. , . 1 13400/.9. .10. 30 Buffer EB . 1 . 11. 1 13400 /. , , , .