Your Materials List
Culture Media
PneumaCult™-NGEx Medium (Catalog #100-1505)
PneumaCult™-NGEx is a defined, serum- and BPE-free cell culture medium for the robust expansion of primary human airway epithelial cells with sustained differentiation potential and function during long-term culture. This medium offers 250x more cells in three weeks—outperforming do-it-yourself (DIY) formulations and other commercially available expansion media.
View Product Details >
PneumaCult™-Ex Plus Medium (Catalog #05040)
PneumaCult™-Ex Plus Medium is a defined, serum- and BPE-free cell culture medium that supports more expansion of primary human airway epithelial cells at each passage, compared to other commercially available expansion media.
PneumaCult™-ALI Medium (Catalog #05001)
PneumaCult™-ALI is a serum- and BPE-free medium for the culture of human airway epithelial cells at the air-liquid interface (ALI). Airway epithelial cells cultured in PneumaCult™-ALI Medium undergo extensive mucociliary differentiation to form a pseudostratified epithelium that exhibits morphological and functional characteristics similar to those of the human airway in vivo.
PneumaCult™-ALI-S Medium (Catalog #05050)
PneumaCult™-ALI-S is a serum- and BPE-free medium for the culture of human small airway epithelial cells at the air-liquid interface (ALI). Small airway epithelial cells cultured in PneumaCult™-ALI-S Medium undergo extensive mucociliary differentiation to form a cuboidal epithelium that exhibits morphological and functional characteristics similar to those of the human small airway in vivo.
PneumaCult™ Airway Organoid Kit (Catalog #05060)
PneumaCult™ Airway Organoid Kit is a serum- and BPE-free culture medium system for efficient establishment and differentiation of airway organoids derived from human bronchial epithelial cells (HBECs). When fully differentiated, the organoids exhibit a centralized lumen surrounded by a polarized airway epithelial cell layer, comprised of differentiated cell types, such as ciliated cells and goblet cells.
PneumaCult™ Apical-Out Airway Organoid Medium (Catalog #100-0620)
Generate reproducible apical-out airway organoids from human bronchial epithelial cells (HBECs) using a Matrigel®-free protocol that enables easy downstream processing.
PneumaCult™ Apical-Out Airway Organoid Secretory Medium (Catalog #100-2078)
PneumaCult™ Apical-Out Airway Organoid Secretory Medium is a serum-, BPE- and Matrigel®-free culture system for the differentiation of airway epithelial cells to mature apical-out organoids containing basal, ciliated, and secretory cells. The fully differentiated organoids are more physiologically relevant making them ideal for downstream applications involving mucus regulation and epithelial diversity.
Transwell® Insert Options for ALI Culture
Costar® 12 mm Transwell®, 0.4 µm Pore Polyester Membrane Inserts (Catalog #38023), or
Costar® 6.5 mm Transwell®, 0.4 µm Pore Polyester Membrane Inserts (Catalog #38024), or
HTS Transwell®-96, 0.4 µm Pore Polyester Membrane Inserts (Catalog #100-0419)
Additional Reagents
Heparin Solution (Catalog #07980)
Hydrocortisone Stock Solution (Catalog #07925)
Animal Component-Free Cell Dissociation Kit (Catalog #05426)
STEMdiff™ Lung Progenitor Kit (Catalog #100-0230)
STEMdiff™ Lung Progenitor Kit is a serum-free culture medium system for efficient and reproducible generation of lung progenitor cells from human embryonic stem (ES) and induced pluripotent stem (iPS) cells through three stages of differentiation: 1) definitive endoderm, 2) anterior foregut endoderm, and 3) lung progenitor cells.
mTeSR™1 (Catalog #85850)
Gentle Cell Dissociation Reagent (Catalog #100-0485)
Y-27632 (Catalog #72302)
STEMdiff™ Branching Lung Organoid Kit (Catalog #100-0195)
STEMdiff™ Branching Lung Organoid Kit (Catalog #100-0195) is a serum-free medium system for efficient and reproducible generation of branching lung organoids from human embryonic stem (ES) and induced pluripotent stem (iPS) cells through four stages of differentiation: 1) definitive endoderm, 2) anterior foregut endoderm, 3) lung bud organoid, and 4) branching lung organoids.
PneumaCult™ Alveolar Organoid Media (Catalog #100-0847)
PneumaCult™ Alveolar Organoid Expansion Medium is a cell culture medium for the passage and long-term expansion of human alveolar epithelial type II (ATII) cells as organoids. The expanded ATII organoids can be differentiated to alveolar epithelial type I (ATI) organoid cultures using PneumaCult™ Alveolar Organoid Differentiation Medium.
24-well non-tissue culture-treated plates (Catalog #100-0097)
Protocol Diagram
Figure 1. Overview of the PneumaCult™ Culture System for Airway Research
Expansion of freshly isolated or commercially available, cryopreserved human bronchial (HBECs), nasal (HNECs), and small airway epithelial cells (HSAECs) in submerged culture can be achieved using PneumaCult™-NGEx. For air-liquid interface (ALI) culture, expanded airway epithelial cells are placed on a cell culture insert. Once confluent, the cells are air-lifted by removing PneumaCult™-NGEx culture medium from both chambers. To differentiate the cells into a mature mucociliary respiratory epithelium, PneumaCult™-ALI (Catalog #05001) is added to the basal chamber for HBECs and HNECs, or PneumaCult™-ALI-S (Catalog #05050) if differentiating HSAECs.
Figure 1. Overview of the PneumaCult™ Culture System for Small Airway Research
Expansion of human small airway epithelial cells (HSAEC) in submerged culture is performed with PneumaCult™-Ex Plus Medium. During the early Expansion Phase of the air-liquid interface (ALI) culture procedure, PneumaCult™-Ex Plus is applied to the apical and basal chambers. Upon reaching confluence, the culture is air-lifted by removing the culture medium from both chambers, and PneumaCult™-ALI-S is added to the basal chamber only. Differentiation into a mucociliary epithelium is obtained following 21+ days of incubation and can be maintained for more than one year.
Figure 1. Overview of the PneumaCult™ Culture System for Human Airway Organoid Generation
In the early two-dimensional expansion phase of the human airway organoid culture procedure, HBECs are expanded using PneumaCult™-NGEx Plus Medium (Catalog # 100-1505). The HBECs are then embedded into a Matrigel® dome and expanded for 4 - 7 days using PneumaCult™ Airway Organoid Seeding Medium. Following the expansion, the HBECs are differentiated using PneumaCult™ Airway Organoid Differentiation Medium for an additional 21+ days.
Figure 1. Generation of Lung Progenitor Cells from Human ES/iPS Cells with the STEMdiff™ Lung Progenitor Kit
Human PSC (hPSC) cultures progress through a simple three-stage process to generate lung progenitor cells. hPSC clumps are first seeded in mTeSR™1. On Day 1, differentiation is initiated with Medium DE-1 (STEMdiff™ Endoderm Basal Medium containing Supplement MR and Supplement CJ). Subsequently on day 2 and 3, the medium is changed to Medium DE-2 (STEMdiff™ Endoderm Basal Medium containing Supplement CJ) for definitive endoderm patterning. On Day 4, to initiate anterior foregut endoderm patterning, the endoderm monolayer is passaged in Medium LP-1 (STEMdiff™ Lung Basal Medium, Lung Supplement (10X), and Supplement 1) and Y-27832. Finally, at Day 7, the cells are differentiated into the lung progenitor stage with Medium LP-2 (STEMdiff™ Lung Basal Medium, Lung Supplement (10X), and Supplement 2).
Figure 1. Generation Of Branching Lung Organoids Using the STEMdiff™ Branching Lung Organoid Kit
(A) Human PSC cultures progress through a four-stage differentiation process to generate human branching lung organoids. By the end of stage 1 (day 3), cultures exhibit characteristics typical of definitive endoderm and anterior foregut differentiation is initiated. During stage 2 (days 3 - 6) anterior foregut endoderm buds are released from the monolayer, and are then suspended to form ventralized lung bud organoids in stage 3 (days 6 - 14). In stage 4, the lung bud organoids are embedded into Matrigel sandwich cultures to mature into branching lung organoids. (B) Morphological representation of the culture at different stages.
Figure 1. Overview of the PneumaCult™ Apical-Out Airway Organoid Secretory Medium Workflow
Prior to generating apical-out airway organoids, human airway epithelial cells (HAECs) are expanded using PneumaCult™-NGEx Medium (Catalog #100-1505) in a two-dimensional expansion phase. The cells are then plated into an AggreWell™400 24-well plate (Catalog #34450) and allowed to aggregate and differentiate using PneumaCult™ Apical-Out Airway Organoid Secretory Medium. Aggregates are cultured in the AggreWell™400 plate for 15 days, after which, fully differentiated organoids are transferred to suspension culture and are immediately assay-ready.
Related Resources
On-Demand Pulmonary Course
Learn from our in-house pulmonary experts as they demonstrate how to culture human airway epithelial cells (HAECs) at the air-liquid interface.
Start Now >
Pulmonary Cell Culture Methods Library
Expansion, maintenance, and differentiation of primary human airway epithelial cells can be challenging. Explore our collection of protocol, tech tips, videos, and more in our Methods Library.
Explore Now >
Have questions about our products or protocols?
Talk to a sales representative about the products in your materials list, recommended protocols, and additional resources and products to support your experimental workflow.