SILVER
SILVER ๋“ฑ๊ธ‰์˜ ํŒ๋งค์ž ์ž๋ฃŒ

[A+]์„œ๊ฐ•๋Œฟ’ํ•™๊ป“ ํ˜„๋Œ€์ƒ๋ฌผํ•™์”„ํ—˜1_4์ฐจ ํ’€๋ ˆํฌํŠธ_ํ•ญ์ฒด ์ƒ์‚ฐ & Western blotting analysis

"[A+]์„œ๊ฐ•๋Œฟ’ํ•™๊ป“ ํ˜„๋Œ€์ƒ๋ฌผํ•™์”„ํ—˜1_4์ฐจ ํ’€๋ ˆํฌํŠธ_ํ•ญ์ฒด ์ƒ์‚ฐ & Western blotting analysis"์— ๋Œ€ํ•œ ๋‚ด์šฉ์ž…๋‹ˆ๋‹ค.
13 ํŽ˜์ด์งฟ’
์›Œ๋“œ
์ตœ์ดˆ๋“ฑ๋ก์ผ 2025.02.23 ์ตœ์ข…์ ฟ’์ž‘์ผ 2024.06
13P ๋ฏธ๋้ฉ๋ณด๊ธฐ
[A+]์„œ๊ฐ•๋Œฟ’ํ•™๊ป“ ํ˜„๋Œ€์ƒ๋ฌผํ•™์”„ํ—˜1_4์ฐจ ํ’€๋ ˆํฌํŠธ_ํ•ญ์ฒด ์ƒ์‚ฐ & Western blotting analysis
  • ๋ฏธ๋้ฉ๋ณด๊ธฐ

    ์†Œ๊ฐœ

    "[A+]์„œ๊ฐ•๋Œฟ’ํ•™๊ป“ ํ˜„๋Œ€์ƒ๋ฌผํ•™์”„ํ—˜1_4์ฐจ ํ’€๋ ˆํฌํŠธ_ํ•ญ์ฒด ์ƒ์‚ฐ & Western blotting analysis"์— ๋Œ€ํ•œ ๋‚ด์šฉ์ž…๋‹ˆ๋‹ค.

    ๋ชฉ์ฐจ

    Abstract
    Introduction
    Materials & Methods
    Result
    Discussion
    References

    ๋ณธ๋ๅฉ๋‚ด์šฉ

    ํ•ญ์›(Antigen)์€ Non-self๋กœ ์ธ์‹๋˜๋Š” ์›์ธ ๋ฌผ์งˆ๋กœ ์ฒด๋‚ด์— ์นจ์ž…ํ•˜๊ณ , ํ•ญ์›์„ ์ธ์‹ํ•œ ๋ฉด์—ญ์„ธํฌ๋Š” ํ•ญ์ฒด๋ฅผ ์ƒ์„ฑํ•œ ๋’ค ์„ธํฌ ์™ธ๋ถ€๋กœ ๋ถ„๋น„ํ•œ๋‹ค. ์ด๋Ÿฌํ•œ ํ•ญ์ฒด๊ฐ€ ํ•ญ์›์˜ Epitope์— ๊ฒฐํ•ฉํ•˜๋ฉฐ ๋ณ‘์›์„ฑ์„ ์ค‘ํ™”์‹œํ‚ค๋ฉฐ ์†Œ๋ฉธ์— ๋„์›€์„ ์ค€๋‹ค. ์ฆ‰, ํ•ญ์ฒด๋Š” ํ•ญ์›๊ณผ ํŠน์ด์ ์œผ๋กœ ๊ฒฐํ•ฉํ•˜๋Š” ํŠน์ง•์„ ๊ฐ–๊ณ  ์žˆ๋‹ค. ์ดํ›„ ๋™์ผํ•œ ํ•ญ์›์ด ์žฌ์นจ์ž…ํ•˜๋ฉด ์ด๋ฅผ ์ธ์‹ํ•œ ๊ธฐ์–ต์„ธํฌ๊ฐ€ ํ˜•์งˆ์„ธํฌ์™ฟ’ ๊ธฐ์–ต์„ธํฌ๋กœ ๋ถ„ํ™”ํ•ด ๋”์šฑ ๋น ๋ฅด๊ฒŒ ํ•ญ์ฒด๊ฐ€ ์ƒ์„ฑ๋œ๋‹ค. ํ•ญ์›์„ ํŠน์ • ๋™๋ฌผ์— ์ฃผ์‚ฌํ•˜์—ฌ ํ•ญ์ฒด ํ˜•์„ฑ์„ ์œ ๋„ํ•  ์ˆ˜ ์žˆ์œผ๋ฉฐ, ๋ฉด์—ญ ๋ฐ˜์‘์„ ์ด‰์ง„ํ•˜๋Š” Adjuvant๋ฅผ ์‚ฌ์šฉํ•œ๋‹ค. ์ถ”๊ฐ€ ์ ‘์ข…์€ ์ฒซ ์ ‘์ข… ์ดํ›„ ์ตœ์†Œ 7์ผ ์ดํ›„์— ์‹ค์‹œํ•˜๋ฉฐ, ์ถ”๊ฐ€ ์ ‘์ข… ํ›„ 3-4์ฃผ ํ›„์— ํ˜ˆ์žฅ์˜ ํ•ญ์ฒด ์–‘์ด ์ตœ๋Œ€์น˜๊ฐ€ ๋œ๋‹ค. ํ•ญ์ฒด ์ƒ์‚ฐ์—๋Š” ์ฃผ๋กœ ์ฅ(Mus musculus)๋ฅผ ์‚ฌ์šฉํ•˜๊ณ , ์ฑ„ํ˜ˆ ์‹œ ์‹ฌ์žฅ ์ฑ„ํ˜ˆ์€ ๋™๋ฌผ์—๊ฒŒ ๊ณ ํ†ต์„ ๋™๋ฐ˜ํ•  ์ˆ˜ ์žˆ์–ด ์•ˆ๋ฝ์‚ฌ๋ฅผ ์ง„ํ–‰ํ•œ ๋‹ค์Œ ํ˜ˆ์•ก์„ ์ฑ„์ทจํ•œ๋‹ค. SDS-PAGE๋ฅผ ํ†ตํ•ด (-) ์ „ํ•˜๋ฅผ ๋„๋Š” ๋‹จ๋ฐฑ์งˆ๋“ค์„ ์˜ค๋กœ์ง€ ๋ถ„์ž๋Ÿ‰์— ๋”ฐ๋ผ ๋‹จ๋ฐฑ์งˆ์„ ๋ถ„๋ฆฌํ•  ์ˆ˜ ์žˆ๊ณ , Immunoblotting์œผ๋กœ ๋ถ„๋ฆฌํ•œ ๋‹จ๋ฐฑ์งˆ์— specificํ•œ ํ•ญ์ฒด๋ฅผ ์ด์šฉํ•˜์—ฌ ํŠน์ • ๋‹จ๋ฐฑ์งˆ์˜ ์œ ๋ฌด๋ฅผ ํ™•์ธํ•œ๋‹ค. 6x His tagged eGFP์™ฟ’ Adjuvant๋ฅผ 1:1 ๋น„์œจ๋กœ ์„ž์€ mixture๋ฅผ ํ•ญ์›์œผ๋กœ ์ผ์ฃผ์ผ ๋‹น 150ul ์ด 3ํšŒ ์ ‘์ข…ํ•œ mouse์—์„œ ํ•ญ์ฒด๋ฅผ ์ƒ์‚ฐํ•˜์˜€์œผ๋ฉฐ, 4์ฃผ์ฐจ์— ํ˜ˆ์•ก์„ ์ฑ„์ทจํ•˜์—ฌ eGFP์— ๋Œ€ํ•œ ํ•ญ์ฒด๊ฐ€ ์กด์žฌํ•˜๋Š” ํ˜ˆ์ฒญ์„ ์–ป์—ˆ๋‹ค. ์ดํ›„, GFP 1x (0.5mg/ml), 1/2 x, 1/4 x, 1/8 x sample์„ SDS-PAGE๋ฅผ ์‹ค์‹œํ•˜๊ณ  ์ดํ›„ membrane์— transfer ํ›„ mouse์—์„œ ์–ป์€ ํ•ญ์ฒด๋ฅผ 1์ฐจ๋กœ ์ด์šฉํ•œ immunoblotting์„ ํ†ตํ•ด ์ตœ์ข…์ ์œผ๋กœ ๋ณด๋ผ์ƒ‰ ๋ฐœ์ƒ‰๋ฐ˜์‘์ด ๋‚˜ํƒ€๋‚œ band๋ฅผ ํ™•์ธํ•˜์˜€๋‹ค.

    ์ฐธ๊ณ ์ž๋ฃŒ

    ยท Oscherwitz, J., Hankenson, F. C., Yu, F., & Cease, K. B. (2006). Low-dose intraperitoneal Freund's adjuvant: Toxicity and immunogenicity in mice using an immunogen targeting amyloid-ฮฒ peptide. Vaccine, 24(15), 3018-3025
    ยท Furie, B., & Furie, B. C. (2008). Mechanisms of thrombus formation. New England Journal of Medicine, 359(9), 938-949
    ยท Sowemimo-Coker, S. O. (2002). Red blood cell hemolysis during processing. Transfusion medicine reviews, 16(1), 46-60
    ยท Gallagher, S. R. (2012). SDSโ€polyacrylamide gel electrophoresis (SDSโ€PAGE). Current Protocols Essential Laboratory Techniques, 6(1), 7-3
    ยท Liu, F., Hashim, N. A., Liu, Y., Abed, M. M., & Li, K. (2011). Progress in the production and modification of PVDF membranes. Journal of membrane science, 375(1-2), 1-27.
    ยท Mahmood, T., & Yang, P. C. (2012). Western blot: technique, theory, and trouble shooting. North American journal of medical sciences, 4(9), 429
    ยท Kitchen, B. J. (1974). A comparison of the properties of membranes isolated from bovine skim milk and cream. Biochimica et Biophysica Acta (BBA)-Biomembranes, 356(3), 257-269
    ยท Cao, Y., Zhao, J., & Xiong, Y. L. (2016). Coomassie Brilliant Blue-binding: a simple and effective method for the determination of water-insoluble protein surface hydrophobicity. Analytical methods, 8(4), 790-795
  • Easy Ai ์š”์•ฝ

    ์ด ๋ฌธ์„œ๋Š” ํ•ญ์ฒด ์ƒ์‚ฐ ๋ฐ Western blotting ๋ถ„์„์— ๋Œ€ํ•œ ๋‚ด์šฉ์„ ๋‹ค๋ฃจ๊ณ  ์žˆ๋‹ค. ํ•ญ์› ์ฃผ์ž…์„ ํ†ตํ•œ ํ•ญ์ฒด ์ƒ์‚ฐ ๊ณผ์ •, SDS-PAGE๋ฅผ ์ด์šฉํ•œ ๋‹จ๋ฐฑ์งˆ ๋ถ„๋ฆฌ, Immunoblotting์„ ํ†ตํ•œ ํŠน์ • ๋‹จ๋ฐฑ์งˆ ๊ฒ€์ถœ ๋“ฑ ์ผ๋ จ์˜ ์‹คํ—˜ ๊ณผ์ •๊ณผ ๊ฒฐ๊ณผ๊ฐ€ ์ž์„ธํžˆ ์„ค๋ช…๋˜์–ด ์žˆ๋‹ค. ํ•ญ์ฒด ์ƒ์‚ฐ์—๋Š” ์ฃผ๋กœ ์ฅ๊ฐ€ ์‚ฌ์šฉ๋˜๋ฉฐ, ํ•ญ์ฒด ์ƒ์‚ฐ์„ ์œ„ํ•ด Freund's Adjuvant๋ฅผ ์‚ฌ์šฉํ•˜์—ฌ ๋ฉด์—ญ ๋ฐ˜์‘์„ ์ฆ๊ฐ•์‹œํ‚จ๋‹ค. SDS-PAGE๋ฅผ ํ†ตํ•ด ๋‹จ๋ฐฑ์งˆ์„ ํฌ๊ธฐ์— ๋”ฐ๋ผ ๋ถ„๋ฆฌํ•˜๊ณ , Immunoblotting์œผ๋กœ ํŠน์ • ๋‹จ๋ฐฑ์งˆ์„ ๊ฒ€์ถœํ•œ๋‹ค. ๋ณธ ์‹คํ—˜์—์„œ๋Š” 6x His-tagged eGFP ํ•ญ์›์„ ์ฃผ์ž…ํ•˜์—ฌ eGFP์— ๋Œ€ํ•œ ํ•ญ์ฒด๋ฅผ ์ƒ์‚ฐํ•˜๊ณ , Immunoblotting์œผ๋กœ eGFP ๋‹จ๋ฐฑ์งˆ์„ ํ™•์ธํ•˜์˜€๋‹ค. ์‹คํ—˜ ๊ฒฐ๊ณผ, eGFP์— ํŠน์ด์ ์œผ๋กœ ๊ฒฐํ•ฉํ•˜๋Š” ํ•ญ์ฒด๊ฐ€ ์ƒ์„ฑ๋˜์—ˆ์œผ๋ฉฐ, ํฌ์„ ๋†๋„์— ๋”ฐ๋ผ band ๊ฐ•๋„ ์ฐจ์ด๋ฅผ ๋ณด์˜€๋‹ค. ๋‹ค๋งŒ ์ผ๋ถ€ ๋น„ํŠน์ด์  ๊ฒฐํ•ฉ ํ•ญ์ฒด๋กœ ์ธํ•œ ์ถ”๊ฐ€ band๋„ ๊ด€์ฐฐ๋˜์—ˆ๋‹ค. ์‹คํ—˜ ๊ณผ์ •์—์„œ ๋™๋ฌผ ์‹คํ—˜์˜ ์œค๋ฆฌ ์›์น™์ธ 3R ์›์น™์„ ๊ณ ๋ คํ•˜์—ฌ ๋™๋ฌผ ๊ณ ํ†ต์„ ์ตœ์†Œํ™”ํ•˜๋ ค ๋…ธ๋ ฅํ•˜์˜€๋‹ค. ์ „๋ฐ˜์ ์œผ๋กœ ํ•ญ์ฒด ์ƒ์‚ฐ ๋ฐ Western blotting ๋ถ„์„์— ๋Œ€ํ•œ ์ฒด๊ณ„์ ์ด๊ณ  ์ƒ์„ธํ•œ ์‹คํ—˜ ๊ณผ์ •๊ณผ ๊ฒฐ๊ณผ๋ฅผ ์ž˜ ์ •๋ฆฌํ•˜๊ณ  ์žˆ๋‹ค.
  • ์ž๋ฃŒํ›„๊ธฐ

      Ai ๋ฆฌ๋ทฐ
      ํ•ญ์ฒด ์ƒ์‚ฐ ๋ฐ Western blotting ๋ถ„์„์— ๋Œ€ํ•œ ์ฒด๊ณ„์ ์ธ ์‹คํ—˜ ๊ณผ์ •๊ณผ ๊ฒฐ๊ณผ๊ฐ€ ์ž˜ ์ •๋ฆฌ๋˜์–ด ์žˆ๋‹ค.
    • ์ž์ฃผ๋ฌป๋Š”์งˆ๋ๅฉ์˜ ๋‹ต๋ณ€์„ ํ™•์ธํ•ด ์ฃผ์„ธ์š”

      ํ•ดํ”ผ์บ ํผ์Šค FAQ ๋”๋ต–๊ธฐ

      ๊ผญ ์•Œ์•„์ฃผ์„ธ์š”

      • ์ž๋ฃŒ์˜ ์ •๋ณด ๋ฐ ๋‚ด์šฉ์˜ ์ง„์‹ค์„ฑ์— ๋Œ€ํ•˜์—ฌ ํ•ดํ”ผ์บ ํผ์Šค๋Š” ๋ณด์ฆํ•˜์ง€ ์•Š์œผ๋ฉฐ, ํ•ด๋‹น ์ •๋ณด ๋ฐ ๊ฒŒ์‹œ๋ฌผ ์ €์ž‘๊ถŒ๊ณผ ๊ธฐํƒ€ ๋ฒ•์  ์ฑ…์ž„์€ ์ž๋ฃŒ ๋“ฑ๋ก์ž์—๊ฒŒ ์žˆ์Šต๋‹ˆ๋‹ค.
        ์ž๋ฃŒ ๋ฐ ๊ฒŒ์‹œ๋ฌผ ๋‚ด์šฉ์˜ ๋ถˆ๋ฒ•์  ์ด์šฉ, ๋ฌด๋‹จ ์ „์žฌโˆ™๋ฐฐํฌ๋Š” ๊ธˆ์ง€๋˜์–ด ์žˆ์Šต๋‹ˆ๋‹ค.
        ์ €์ž‘๊ถŒ์นจํ•ด, ๋ช…์˜ˆํ›ผ์† ๋“ฑ ๋ถ„์Ÿ ์š”์†Œ ๋ฐœ๊ฒฌ ์‹œ ๊ณ ๊ฐ๋น„๋ฐ”์นด์ง€๋…ธ Viva์˜ ์ €์ž‘๊ถŒ์นจํ•ด ์‹ ๊ณ ๋น„๋ฐ”์นด์ง€๋…ธ Viva๋ฅผ ์ด์šฉํ•ด ์ฃผ์‹œ๊ธฐ ๋ฐ”๋ž๋‹ˆ๋‹ค.
      • ํ•ดํ”ผ์บ ํผ์Šค๋Š” ๊ตฌ๋งค์ž์™ฟ’ ํŒ๋งค์ž ๋ชจ๋‘๊ฐ€ ๋งŒ์กฑํ•˜๋Š” ์„œ๋น„์Šค๊ฐ€ ๋˜๋„๋ก ๋…ธ๋ ฅํ•˜๊ณ  ์žˆ์œผ๋ฉฐ, ์•„๋ž˜์˜ 4๊ฐ€์ง€ ์ž๋ฃŒํ™˜๋ถˆ ์กฐ๊ฑด์„ ๊ผญ ํ™•์ธํ•ด์ฃผ์‹œ๊ธฐ ๋ฐ”๋ž๋‹ˆ๋‹ค.
        ํŒŒ์ผ์˜ค๋ฅ˜ ์ค‘๋ณต์ž๋ฃŒ ์ €์ž‘๊ถŒ ์—†์Œ ์„ค๋ช…๊ณผ ์‹ค์ œ ๋‚ด์šฉ ๋ถˆ์ผ์น˜
        ํŒŒ์ผ์˜ ๋‹ค์šด๋กœ๋“œ๊ฐ€ ์ œ๋Œ€๋กœ ๋˜์ง€ ์•Š๊ฑฐ๋‚˜ ํŒŒ์ผํ˜•์‹์— ๋งž๋Š” ํ”„๋กœ๊ทธ๋žจ์œผ๋กœ ์ •์ƒ ์ž‘๋™ํ•˜์ง€ ์•Š๋Š” ๊ฒฝ์šฐ ๋‹ค๋ฅธ ์ž๋ฃŒ์™ฟ’ 70% ์ด์ƒ ๋‚ด์šฉ์ด ์ผ์น˜ํ•˜๋Š” ๊ฒฝ์šฐ (์ค‘๋ณต์ž„์„ ํ™•์ธํ•  ์ˆ˜ ์žˆ๋Š” ๊ทผ๊ฑฐ ํ•„์š”ํ•จ) ์ธํ„ฐ๋„ท์˜ ๋‹ค๋ฅธ ์‚ฌ์ดํŠธ, ์—ฐ๊ตฌ๊ธฐ๊ด€, ํ•™๊ป“, ์„œ์  ๋“ฑ์˜ ์ž๋ฃŒ๋ฅผ ๋„์šฉํ•œ ๊ฒฝ์šฐ ์ž๋ฃŒ์˜ ์„ค๋ช…๊ณผ ์‹ค์ œ ์ž๋ฃŒ์˜ ๋‚ด์šฉ์ด ์ผ์น˜ํ•˜์ง€ ์•Š๋Š” ๊ฒฝ์šฐ
    ๋ฌธ์„œ ์ดˆ์•ˆ์„ ์ƒ์„ฑํ•ด์ฃผ๋Š” EasyAI
    ์•ˆ๋…•ํ•˜์„ธ์š”. ํ•ดํ”ผ์บ ํผ์Šค์˜ ๋ฐฉ๋Œ€ํ•œ ์ž๋ฃŒ ์ค‘์—์„œ ์„ ๋ณ„ํ•˜์—ฌ ๋‹น์‹ ๋งŒ์˜ ์ดˆ์•ˆ์„ ๋งŒ๋“ค์–ด์ฃผ๋Š” EasyAI ์ž…๋‹ˆ๋‹ค.
    ์ €๋Š” ์•„๋ž˜์™ฟ’ ๊ฐ™์ด ์ž‘์—…์„ ๋„์™ฟ’๋“œ๋ฆฝ๋‹ˆ๋‹ค.
    - ์ฃผ์ œ๋งŒ ์ž…๋ ฅํ•˜๋ฉด ๋ชฉ์ฐจ๋ถ€ํ„ฐ ๋ณธ๋ๅฉ๋‚ด์šฉ๊นŒ์ง€ ์ž๋™ ์ƒ์„ฑํ•ด ๋“œ๋ฆฝ๋‹ˆ๋‹ค.
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