**3. Basal lamina model systems**

A suitable source for the purification of an extracellular matrix that mimics the basal lamina is the Engelbreth–Holm–Swarm sarcoma of mice. This tumor produces, in contrast to healthy tissue, large amounts of ECM with laminin and collagen IV being the main components [3]. Depending on the question asked, individual macromolecular components of the basal lamina may be sufficient to take over the role of the complex biopolymer mixture. For instance, adhesion of cells to solid substrates is promoted similarly well by laminin coatings as by coatings with the multicomponent ECM [40, 41]. For other basal lamina properties such as viscoelasticity and selective permeability, it is crucial that the biological complexity of the system is maintained so that the full spectrum of basal lamina function is obtained.

The high abundance of ECM in Engelbreth–Holm–Swarm tumor tissue makes it possible to purify reasonable amounts of this multicomponent matrix as required for systematic *in vitro* experiments. A first purification protocol for this ECM was established by Kleinman et al. [42, 43] in the 1980s. The extract is liquid at temperatures between 4°C and approx. 15°C and forms a gel at higher temperatures. In its gel form, the matrix was tested for its biological activity, and it was shown in several studies that the purified ECM successfully promotes the differ‐ entiation of various cell types [44–47]. Cells can be either plated on top of the gel, thus simulating a two-dimensional (2D) environment, or they can be embedded into a 3D ECM matrix. Which configuration is chosen depends on the detailed experimental setup, the cell type used and the biological question. For instance, cell migration experiments can be con‐ ducted both on flat surfaces which have been coated by ECM components and in 3-dimensional basal lamina gels [48, 49].

The purification protocol of Kleinman et al. is used by several companies for the commercial production of ECM. Although these commercial ECM variants are extracted according to the same purification protocol, significant differences in the behavior of cells embedded into those gels have recently been described [50]: The migration behavior of leukocyte-like dHL-60 cells in four different commercially available ECM gel variants differed strongly even though the gels were prepared at matching total protein concentrations. Moreover, in one of the ECM gels, life–dead stains demonstrated a significantly increased percentage of nonviable cells. At the same time, for this gel variant, there was an additional band visible when the gel was analyzed by SDS-PAGE. Mass spectrometry showed that this additional band contained laminin fragments which indeed are suspected to be harmful for cells. This result demonstrates the dilemma a researcher is exposed to when working with commercial model systems: On the one hand, the relatively easy availability of the material in reasonable quantities allows for conducting *in vitro* experiments which otherwise would not be possible. On the other hand, comparability to results from other researchers is often difficult if different vendor sources for the biopolymer mixture are used: The data obtained need to be interpreted with great care and ideally should be double-checked with a second, independent ECM preparation. SDS-PAGE analysis also suggested that the commercial ECM variants differed in terms of the relative concentration of basal lamina components. Whereas, in all ECM preparations, the bands corresponding to collagen IV and laminin were clearly most pronounced, the strongest variability occurred in a band around 50 kDa which matches the molecular weight of entactin. Since this molecule acts as a cross-linker between laminin and collagen IV, it is reasonable to assume that variations in its relative concentration will also affect the structure and permea‐ bility properties of the ECM gels and, ultimately, the migration behavior of cells in those gels. However, biochemical techniques are not able to predict those gel parameters, which is why physical methods are required to further characterize the different basal lamina model systems.
