• NanoString共催!ウェビナー オンデマンド配信開始! GeoMxによる空間解析の可能性
  • 技術情報特集ページ 空間プロファイリング解析 GeoMx解析サービスの特集記事連載中
  • 画像解析ソフトウェア HALOによる画像解析 スコアリング・数値化
ホーム >  技術情報  >  免疫組織化学画像(抗体リスト)  > 文献紹介





 下記の論文では、変異EGFRに対する抗体を作製し、その評価を行っています。EGFR変異のおよそ90%を占めると言われている2つの変異(exon19 del.、L858R変異)に対する抗体を作製し、あらかじめEGFR変異の情報が明らかな培養細胞やゼノグラフト組織サンプルを用いたウェスタンブロット、免疫組織蛍光染色、IHCによってその特異性を確認しました。さらにNSCLCの腫瘍組織のEGFR変異について、IHC法、シークエンス法およびMS-basedシークエンス法の解析結果を比較したところ、IHCで92%の感度と99%の特異性を示し、さらにシークエンス法よりも精度が高いことが示唆されました。今後、この抗体を用いた検査によってEGFR-TKIの奏功性予測検査が迅速・簡便・低コストで可能になることが期待できます。

Mutation-Specific Antibodies for the Detection of EGFR Mutations in Non-Small-Cell Lung Cancer
Yu J, Kane S, Wu J, Benedettini E, Li D, Reeves C, Innocenti G, Wetzel R, Crosby K, Becker A, Ferrante M, Cheung WC, Hong X, Chirieac LR, Sholl LM, Haack H, Smith BL, Polakiewicz RD, Tan Y, Gu TL, Loda M, Zhou X, Comb MJ.
Cancer Res 2009;15 : 3023-3028


Purpose: Activating mutations within the tyrosine kinase domain of epidermal growth factor receptor (EGFR) are found in approximately10% to 20% of non-small-cell lung cancer (NSCLC) patients and are associated with response to EGFR inhibitors. The most common NSCLC associated EGFR mutations are deletions in exon 19 and L858R mutation in exon 21, together accounting for 90% of EGFR mutations. To develop a simple, sensitive, and reliable clinical assay forth e identification of EGFR mutations in NSCLC patients, we generated mutation-specific rabbit monoclonal antibodies against each of these two most common EGFR mutations and aimed to evaluate the detection of EGFR mutations in NSCLC patients by immunohistochemistry.
Experimental Design: We tested mutation-specific antibodies by Westernblot, immunofluorescence, and immunohistochemistry. In addition, we stained 40 EGFR genotyped NSCLC tumor samples by immunohistochemistry with these antibodies. Finally, with a panel of four antibodies, we screened a large set of NSCLC patient samples with unknown genotype and confirmed the immunohistochemistry results by DNA sequencing.
Results: These two antibodies specifically detect the corresponding mutant form of EGFR by Western blotting, immunofluorescence, and immunohistochemistry. Screening a panel of 340 paraffin-embedded NSCLC tumor samples with these antibodies showed that the sensitivity of the immunohistochemistry assay is 92%, with a specificity of 99% as compared with direct and mass spectrometry-based DNA sequencing.
Conclusions: This simple assay for detection of EGFR mutations in diagnostic human tissues provides a rapid, sensitive, specific, and cost-effective method to identify lung cancer patients responsive to EGFR-based therapies.

PMID 19366827



Assessment of EGFR mutation status in lung adenocarcinoma by immunohistochemistry using antibodies specific to the two major forms of mutant EGFR
Marie Brevet, Maria Arcila, Marc Ladanyi
J Mol Diagn. 2010 Mar;12(2):169-76               


 EGFR mutations are the best predictors of response to EGFR kinase inhibitors in lung adenocarcinoma. We evaluated two mutation-specific monoclonal antibodies for the detection of EGFR mutations by immunohistochemistry (IHC), generated respectively against the L858R mutant and the exon 19 mutant with the common 15bp/5AA deletion. These two mutations account for approximately 90% of all EGFR mutations. IHC staining performed on 218 paraffin-embedded lung adenocarcinomas was assessed on a 0 to 3+ scale, and positivity cutoffs of 1+ and 2+ were compared. All cases were studied by standard molecular methods for these two mutations, and selected cases were also studied using higher sensitivity molecular assays. The EGFR L858R mutant antibody showed a sensitivity of 95% and a positive predictive value (PPV) of 99% with a positivity cutoff of 1+ and a sensitivity of 76% and a PPV of 100% with a positivity cutoff of 2+. The EGFR exon 19 mutant-specific antibody showed reduced sensitivity for exon 19 deletions other than 15bp. A positivity cutoff of 1+ resulted in a sensitivity of 85% and a PPV of 99%, whereas a 2+ cutoff gave a sensitivity of 67% and a PPV of 100%. IHC with EGFR mutant-specific antibodies could be used as a screen to identify most candidates for EGFR inhibitors.

PMID 20093391