HistoGeneX is now CellCarta Learn More

PD-L1 (22C3) status (Lung Cancer)


To determine the PD-L1 status in non-small cell lung cancer (NSCLC), the PharmDx PD-L1 22C3 IHC test from Agilent is used. This Dako 22C3 PharmDx IHC assay is performed as a fully automated immunohistochemistry assay on the Dako autostainer Link 48 automated slide stainer.

Clinical Implication

On October 24 2016, the FDA announced that KEYTRUDA (pembrolizumab) is now approved for the first-line treatment of metastatic NSCLC patients whose tumors express high levels of PD-L1 (Tumor Proportion Score [TPS] of 50 percent or more) or for previously treated metastatic NSCLC patients whose tumors express PD-L1 (TPS of 1 percent or more). PD-L1 expression levels are determined using the 22C3 PharmDx assay (1). Prior to this, chemotherapy was the standard first-line treatment for most NSCLC patients. Pembrolizumab is the second FDA-approved PD-1 inhibitor in lung cancer, and first across all NSCLC histologies. The PD-1 agent nivolumab (Opdivo) was approved in March 2015 for patients with NSCLC who have progressed on or after platinum-based chemotherapy. However, the indication is limited to individuals with squamous histology.

A total of ~6000 patients were screened for eligibility in clinical trials KEYNOTE-001, KEYNOTE-010, and KEYNOTE-024. This is one of the largest data sets of PD-L1 expression determined by an FDA-approved companion diagnostic in patients with advanced NSCLC screened for pembrolizumab therapy. 68% of patients with advanced NSCLC had PD-L1 TPS ≥1% and 28% had PD-L1 TPS ≥50%. The prevalence is similar across prior lines of therapy and different disease characteristics examined (3,4).

Specimen Requirements

Acceptable specimens for the assay are formalin (10% buffered or Zinc formalin)-fixed (6-48 hours), paraffin-embedded NSCLC tissue specimens. Sectioned slides must be analysed within 3 months after sectioning date. 

In archival tissue of > 6 months old the PD-L1 protein can be deteriorated resulting in unreliable staining (3).


1 representative paraffin block is preferred. Alternatively, 3 unstained freshly sectioned tissue sections are accepted (1 slide of 4 or 5 µm thickness for H&E staining and a minimum of 2 slides of 4-5 µm thickness for PDL1 IHC testing).

Storage and Shipment Instructions

Maintain and ship specimens at ambient temperature.


Fixatives other than buffered formalin, prolonged fixation time, decalcification methods or exposure to severe heat or chemicals may give rise to inadequate results.

Special Requirements


Turn-Around Time

Three to 5 business days for slides and paraffin blocks respectively.


  1. https://www.agilent.com/about/newsroom/presrel/2016/24oct-ca16033.html
  2. Gandhi L, Balmanoukian A, Hui R, et al. Abstract CT105: MK-3475 (anti-PD-1 monoclonal antibody) for non-small cell lung cancer (NSCLC): Antitumor activity and association with tumor PD-L1 expression. Cancer Research. 2014;74:CT105
  3. Garon EB et al. Pembrolizumab for the treatment of non-small-cell lung cancer. N Engl J Med. 2015;372:2018–28
  4. Grigg C et al. PD-L1 biomarker testing for non-small cell lung cancer: truth or fiction? J Immunother Cancer. 2016; 4: 48.

Updated on 09 April 2019