What is a U-ULA Plate?

Ultra-Low Attachment (ULA) plates are specialized cell culture plates designed to prevent cell adhesion, allowing cells to self-aggregate into spheroids in suspension. U-Bottom Ultra-Low Attachment (U-ULA) plates combine the benefits of a U-bottom well shape with an ultra-low attachment surface, ensuring that cells settle centrally and form consistent spheroids without adhering to the plate. These plates are widely used in drug discovery, cancer research, and high-throughput screening. 

How Do ULA Plates Work?

• ULA plates are coated with an ultra-low attachment surface, preventing cells from adhering to the well.
• Cells naturally aggregate in 3D spheroids, mimicking in vivo tumor conditions.
• Used for high-throughput drug screening, toxicity testing, and tissue engineering.

Why Scaffold-Free U-ULA Plates are Important in HTS/HCS Applications

Scaffold-free cell culture technologies, such as U-ULA plates, play a crucial role in high-throughput screening (HTS) and high-content screening (HCS) due to their simplicity, cost-effectiveness, and compatibility with automation. These advantages make them an essential tool for researchers conducting large-scale drug discovery and cellular studies.

Ease of Use & Cost-Effectiveness

One of the primary benefits of U-ULA plates is their ease of use and cost-effectiveness. Unlike scaffold-based systems that require additional materials such as hydrogels or extracellular matrix components, U-ULA plates allow cells to naturally aggregate without external support. This reduces material costs and simplifies the experimental workflow, making them an ideal choice for large-scale studies. Additionally, because U-ULA plates are pre-coated and ready to use, researchers can bypass extensive preparation steps, saving both time and resources.

High-Throughput Compatibility

U-ULA plates are designed for high-throughput applications, making them fully compatible with automated liquid handling systems. This allows for efficient dispensing, culturing, and screening of cells in multi-well plate formats. With availability in 96-well and 384-well configurations, U-ULA plates enable researchers to conduct parallel testing of thousands of drug compounds simultaneously. This scalability is crucial in pharmaceutical research, where screening large compound libraries efficiently is essential for accelerating drug discovery.

Controlled & Consistent Spheroid Formation

One of the significant challenges in 3D cell culture is ensuring uniform spheroid formation across all wells. U-ULA plates are designed to generate a single spheroid per well, ensuring consistency and reproducibility in experimental outcomes. This controlled environment is particularly important in HTS, where standardization is required for reliable comparisons across different drug treatments. By reducing variability in spheroid size and structure, U-ULA plates improve the accuracy of screening assays and minimize data inconsistencies.

Improved Imaging & Analysis

The ability to accurately track and analyze spheroid development is critical in high-content imaging applications. U-ULA plates facilitate straightforward imaging because spheroids can be typically found in a predictable location. Furthermore, U-ULA plates help reduce background noise in fluorescence and confocal microscopy, leading to clearer imaging results. This feature is especially beneficial for researchers conducting time-lapse imaging, live-cell tracking, and multi-channel fluorescence analysis.

U-ULA plates provide a reproducible, cost-effective, and scalable solution for high-throughput and high-content screening applications. Their ability to support consistent spheroid formation, compatibility with automation, and enhanced imaging capabilities makes them an essential tool for drug discovery and biomedical research

Why Compare U-ULA Plates to GrowDex?

Limitations of U-ULA Plates 

1. Inefficient Spheroid Formation for Some Cell Types: Not all cell types efficiently form spheroids in U-ULA plates. Instead, these cells often remain in loose aggregates, leading to irregular morphology and inconsistent experimental results. This limitation is particularly problematic for researchers aiming to develop standardized 3D culture models.
2. Imaging Challenges Due to Spheroid Movement: In HCS and fluorescence imaging, spheroid stability is crucial for obtaining clear, high-resolution images. One major drawback of U-ULA plates is that spheroids tend to roll or shift, making it difficult to capture images from a consistent focal plane. This movement can introduce blur and inconsistencies in fluorescence signal detection, complicating multi-channel imaging and time-lapse studies. In contrast, hydrogel-based systems like GrowDex provide a stable environment that immobilizes spheroids, ensuring precise imaging over time.
3. Lack of Cell-Matrix interactions: The extracellular matrix (ECM) is crucial for mimicking physiological cell-matrix interactions. Without ECM, spheroids rely solely on cell-cell adhesion, leading to weaker structural integrity, inaccurate tumor microenvironment modeling, and altered drug responses. ECM also supports cell migration and invasion, which U-ULA plates cannot replicate. This limitation affects tumor modeling and therapy testing, making ECM-based models more biologically relevant for cancer research and personalized medicine.
4. Lower Cell Viability Over Extended Culture Periods: Short term cultures and treatments using U-ULA spheroids can work well, however for long-term cell culture experiments, spheroids generated in U-ULA plates may develop a necrotic core, and show lower viability compared to hydrogel-based scaffolds. This makes ULA plates less suitable for extended studies, particularly for research involving long-term drug exposure or tissue regeneration models.

While U-ULA plates are useful for high-throughput spheroid culture, their limitations in morphology control, reproducibility, imaging, and long-term viability make them less ideal for advanced 3D cell culture applications. Researchers seeking consistent, physiologically relevant models may benefit from hydrogel-based alternatives like GrowDex, which offer better spheroid formation, stability, and viability.

U-ULA Plates Vs GrowDex : A Comparison Study

GrowDex, a nanocellulose-based hydrogel, provides a 3D ECM like structure that enhances spheroid formation, maintains high viability, and improves imaging quality. Therefore, a comparison study was performed in house at UPM Biomedicals aiming to evaluate spheroid formation, viability, and reproducibility in U-bottom ULA plates versus GrowDex/GrowDex-T hydrogels across multiple cancer and normal cell lines.

Key Questions Answered in This Study:

1. How does spheroid morphology compare between U-ULA plates and GrowDex?
2. Do cells remain viable for longer in GrowDex?
3. Which system provides more reproducible results across different experiments?
4. How does imaging quality differ between the two systems?

Cell Line Performance: GrowDex vs. U-ULA

This study classified spheroids using the National Cancer Institute’s NCI60 categorization (Selby et al. 2017) based on the following categorisation:

• Category 1: Condensed spheroids
• Category 2: Spheroidal shapes 
• Category 3: Aggregates
• Category 4: Loose aggregates

Results from the Study

1. Spheroid Morphology & Formation

The study demonstrated that GrowDex/GrowDex-T effectively supports compact spheroid formation, even in challenging cell types that failed to form spheroids in U-ULA plates. The natural nanocellulose matrix in GrowDex creates a more structured environment, leading to better cell-cell interactions and uniform spheroid growth. In contrast, U-ULA plates often resulted in loose aggregates, with spheroids displaying irregular morphology. GrowDex/T cultures also exhibited higher reproducibility, ensuring consistent spheroid formation across experiments, which is a crucial factor in drug discovery and cell-based screening.

2. Viability & Longevity

Cell viability was significantly higher in GrowDex cultures, particularly in long-term experiments lasting 7–8 days. The hydrogel's extracellular matrix like properties provided structural and biochemical support that enhanced cell survival. In contrast, U-ULA cultures exhibited lower viability, with some cell lines showing early cell death. Notably, mIMCD-3 kidney cells died by Day 4 in U-ULA plates, while they remained viable and proliferative in GrowDex/GrowDex-T cultures. Additionally, some cell types, such as PANC-1 and PC3, showed better spheroid morphology and growth in GrowDex-T compared to standard GrowDex, highlighting the importance of hydrogel selection based on cell type.

3. Imaging & High-Content Screening (HCS) Considerations

For HCS and fluorescence microscopy, GrowDex performed better than U-ULA plates. Using flat-bottom (F-bottom) plates with GrowDex resulted in better imaging clarity, as it provided a stable matrix that prevented spheroid movement. Conversely, U-bottom ULA plates had a small imaging window, making it difficult to capture high-resolution images. In live-cell imaging and fluorescence-based assays, spheroids in ULA plates tended to roll or shift, affecting the precision of time-lapse studies. By contrast, GrowDex immobilized spheroids, ensuring consistent positioning for repeated imaging and analysis, that is an essential feature for HCS workflows.

4. Suitability for Different Cell Lines

The study revealed that GrowDex outperformed U-ULA plates in most tested cell lines, particularly in cancer and kidney-derived cells. While some cell lines, such as A549 lung cancer cells, showed similar morphology in both platforms, others, like MCF7 (breast cancer), PC3 (prostate cancer), and IGROV-1 (ovarian cancer), exhibited more compact and well-formed spheroids in GrowDex. Additionally, mIMCD-3 kidney cells struggled to survive in U-ULA plates, reinforcing that GrowDex provides superior support for long-term cultures.

Summary from the Study: 

Results showed that MCF7, PC3, and IGROV-1 cells formed compact spheroids in GrowDex but remained loose aggregates in U-ULA plates. mIMCD-3 (kidney cells) had higher viability in GrowDex, whereas cells died early in U-ULA plates. Imaging fluorescent-labeled spheroids was more effective in GrowDex, as it immobilized spheroids, preventing movement.

Since GrowDex consistently maintains viability, reproducibility, and better imaging, we can use it as:

• Complementary tool to U-ULA plates for users looking for better spheroid morphology and viability.
• A superior alternative to U-ULA plates in cases where ULA plates fail to form consistent spheroids.
• Ideal for high content imaging applications, offering better control over spheroid positioning.

GrowDex: A Powerful Complement to U-ULA Plates

While U-ULA plates are useful for high-throughput spheroid screening, they do not work for all cell types and often lack a controlled ECM-like environment. Additionally, there can be imaging and analysis issues due to movement of the spheroids within the U-bottom well, in response to fast plate movement during high throughput screening.

GrowDex is a natural nanocellulose hydrogel that provides better spheroid formation, higher cell viability, and improved imaging for 3D cell culture applications. GrowDex hydrogels can be used to mechanically inhibit spheroid movement within the well during high throughput screening.

When to Choose GrowDex vs. U-ULA Plates

References:

3D Models of the NCI60 Cell Lines for Screening Oncology Compounds - Selby, Mike et al. SLAS Discovery, Volume 22, Issue 5, 473 - 483