WITH FOCUS ON LIVER&ISLETS
PELOBiotech Newsletter June/July 2019
“The heart has the nature of
knowledge, the liver of emotion,
the lungs of leaf..“
the liver plays a critical role in human metabolism. As the gatekeeper of the digestive
track, this huge organ is responsible for drug breakdown and is therefor
the first to be injured due to overdose or misuse. Evaluating this drug-induced
liver injury is a critical part of pharmaceutical drug discovery and must be carried
out on human liver cells.
Pancreatic cancer is one of the most challenging research fields
today. Cancer develops when cells grow in an uncontrolled manner
and form masses or tumors in the pancreas. Due to the importance
of these organs, we have assembled this issue. Enjoy!
Editor in Chief PELONews
New Sales Contact in France
—Hildegard von Bingen,
German Nun & Healer-
Liver/Pancreas 2D/3D models
Enzymes: Revolution of
for Human Islet Isolation
Reducing the Cost of Human
Especially for our French customers we have
an own dependance now for you in France.
NEW: 2D/3D Hepatocyte
I like to introduce to you M. Michel Vidal. He´s
representing our products and service to you and
very happy to talk to you about your research projects
M. Vidal is very experienced in the Life Science
sector and can support you in your daily business.
Please contact us or your personal contact in France
M. Michel Vidal : Mobile + 33 (0)6 07 75 16 53
DNs-Rh: fluorogenic substrate
Trabecular Meshwork Cells
You like what you see? Sign
up for PELONews now at
www.pelobiotech.com or just
fill out the following form
Liver/Pancreas 2D/3D models
Go natural: Take native Hydrogels
Xylyx Bio has developed the native proteins and growth factors from the respective organ tissue for the cell culture and
thus reproduce a more natural environment. The ECM is available as coating, hydrogel or scaffold (custom order) for liver,
pancreas-and much, much more.
Pancreas: 2D Surface Coating , 3D Scaffold
Liver: 2D Surface Coating , 3D Hydrogel & Scaffold
Take your pick: Pancreas ECM cell culture substrates Exxtracellular matrix (ECM) is a major component of the cell microenvironment
with important signaling and regulatory functions. Derived from healthy acellular porcine porcine tissue, NativeCoat and
TissueSpec® pancreas matrix is rich in collagen types I, III, IV but also includes collagen type V.
TissueSpec® liver matrix is the organizational framework consisting of fibronectin and collagen types I, III, IV, V, and VI that mediates
hepatic endothelial and epithelial cell communication, modulates liver homeostasis, and supports repair and regeneration.
Excellent Growth Results for
Dog Primary Liver Organoids
The very innovative company Noviocell has developed Noviogel.
It is a synthetic matrix, free of animal and undefined components.
The hydrogel for cell culture behaves similar to collagen
or fibrin: Your Plus:
- functionalized with RGD peptides or Pure
- thermosensitive and reversible
But look for yourself! Already cultured successfully: Dog primary
liver organoids with Noviogel-RGD: Dog primary liver cells
were cultured in Matrigel and noviogel-RGD for 7 days. Liver
organoids cultured in noviogel-RGD showed excellent growth
characteristics, more organoids and bigger in size.
3D Co Culture Model for Liver, Pancreas Tumor Cells & more
Have a look at theese Applicable Cell Systems for Pancreas and Liver Tumor
cells. These cells have been tested sucessfuly for aggregate sphere formation
by our academic partners.
Full overview here: https://biopply.com/assets/Uploads/Applicable-Cell-
Details are described in the publication: A deep conical agarose microwell array for adhesion independent threedimensional
cell culture and dynamic volume measurement Andreas R. Thomsen et al., Lab Chip, DOI 10.1039/
Revolution of Enzyme Requirements for
Human Islet Isolation by Bob McCarthy
The quality of collagenase used for mammalian islet isolation has been a focus of clinical
research scientists soon after Ballinger and Lacy demonstrated that islet transplantation
could correct chemically induced diabetes in inbred strains of rats
(1). Clostridium histolyticum collagenase became a critical reagent because of the need
to isolate sufficient numbers of functional islets for islet transplantation in higher
mammals (e.g., dogs, pigs, humans). Typically, researchers evaluated several “lots” of
crude or enriched commercial collagenase products (i.e., traditional collagenase) for
their ability to recover pancreatic islets. The goal of lot qualification was to identify a
“good” lot of traditional collagenase that worked successfully in the islet isolation procedure
before purchasing larger amounts of a specific lot of product for routine use.
Good lots for human islet isolation were hard to find.
Camillo Ricordi’s development of a semi-automated method for human islet isolation
led him to focus on screening new collagenase lots once these lots were released for
sale by a supplier. During this process, Ricordi screened lots of Collagenase P, a crude
collagenase product first sold by Boehringer Mannheim Biochemicals in 1990. He
found that this material worked better for this application than competitive collagenase
products. One lot, lot 9, was used to isolate islets for many allo-islet transplantation
procedures (2). Collagenase P had higher collagenase and lower protease activity than
found in Sigma collagenase products used by many research scientists at that time.
Ricordi’s use of the Collagenase P product in clinical trials led to a meeting with R&D
management at Boehringer Mannheim Diagnostics in Indianapolis, Indiana about Boehringer
solving the “collagenase problem.” This discussion led to funding of a research
project at Boehringer Mannheim Biochemicals to identify and purify the key enzymes in
crude collagenase responsible for releasing islets from porcine or human pancreatic
tissue. The assumption was that the identification, purification, and formulation of purified
enzymes used at an appropriate dose would resolve the collagenase problem.
To read full paper, please visit: https://www.vitacyte.com/news/evolution-enzymerequirements-human-islet-isolation-link-recent-review/
1 Ballinger WF, Lacy PE. Transplantation of intact pancreatic islets in rats. Surgery. 1972;72(2):175-86.
2 Ricordi C, Tzakis AG, Carroll PB, Zeng YJ, Rilo HL, Alejandro R, et al. Human islet isolation and allotransplantation
in 22 consecutive cases. Transplantation. 1992;53(2):407-14.
VitaCyte’s defined enriched
products set a new
standard of consistency
and enables assessment
of a broader range of
ratios for cell isolation
than found with current
DE Collagenase is
designed with increasing
amounts of enriched
to a fix amount of purified
Islet Overview discussion
protocols used in rodent
Includes a link to
a white paper that
summarizes a survey
of the affect of rodent
strain, age, sex, or
nutritional state on
expected islet yields
when purified enzymes
were used to isolate
Reducing the Cost of Human Islet Isolation
Clostridium histolyticum collagenase is a critical reagent for isolating islets from human pancreatic tissue. The development
of Liberaseä HI Purified Enzyme Blend overcame the problem of repetitive evaluation of different crude
collagenase lots to identify an acceptable product. The potential threat of transmission of spongiform encephalopathy
from bovine tissue led NIH to disallow the use of Liberase HI in the Clinical Islet Transplantation Consortium
(CITC) trial. Initially, Nordmark/Serva was chosen as the sole enzyme vendor for this trial, but VitaCyte and Roche
were added as suppliers after several experienced islet transplant laboratories were unable to achieve human islet
yields comparable to those obtained when using Liberase HI. All of the enzyme formulations in the CITC trial used
about 20 Wünsch Units of collagenase per g of trimmed tissue, the same dose used in the Immune Tolerance Network
Trial for human islet transplantation. Human islet isolation is an inefficient process since ≈ 53% of the islet isolations
had a sufficient number of islets for subsequent transplantation. The use of more consistent collagenase reagents
may increase the efficiency of this process.
A new era of understanding enzyme-mediated tissue dissociation began when Matsushita’s laboratory reported the
gene sequence and protein domain structure of C. histolyticumcollagenase. His laboratory showed class I (C1) and
class II (C2) collagenase were expressed by separate genes, validating earlier observations that showed these two
classes had different enzyme activity profiles and were separated by chromatographic techniques. They also showed
the degradation of native collagen only occurred with those molecular forms of collagenase that contained a catalytic
domain and at least one collagen binding domain.
VitaCyte correlated the specific collagen degradation activity (CDA) of purified natural collagenase to the molecular
form of the enzyme. Intact C1 with two collagen binding domains was 7-10% more active in degrading native collagen
when compared to “truncated” C1 or intact C2, each having only one collagen binding domain.
The new information on collagenase structure-function led to the development of a hypothetical model for collagenase-protease
digestion of tissue. The six key principles from this model are:
1. Differences in the efficiency of intact C1 vs truncated C1 or intact C2 likely reflects the ability of intact C1 with
two collagen binding domains to stick tighter to the collagen monomer during the degradation process than those
forms with a single collagen binding domain.
2. Collagenase appears to degrade collagen by moving from the amino terminal end of the collagen monomer to
carboxy terminal, degrading a collagen monomer in a fiber or fibril.
3. Excess collagenase CDA must be used to ensure rapid degradation of collagen; excess purified collagenase will
have a minimal adverse effect on cells since it has a narrow specificity for native collagen and gelatin.
4. Collagenase products that contain predominantly intact collagenase will have the highest specific CDA, and can
be used at lower doses in the islet isolation procedure than those products that contain multiple forms of
5. The ratio of C1 to C2 is unlikely to impact the efficiency of degradation of collagen in the extracellular matrix
since these two enzymes work cooperatively to degrade native collagen.
6. Choice and dose of neutral protease used for the isolation procedure is the critical variable that must be controlled
if excess intact collagenase is used in the isolation procedure.
Read full paper here: https://www.vitacyte.com/news/reducing-cost-human-islet-isolation
Read here common protocols used in human islet isolation.
Includes a table listing factors to consider when focusing
on enzymes used in these isolation procedures:
Rodent Hepatocytes Overview. Overview
discussion of methods for isolating rodent
hepatocytes. Includes a table of recommended
products and doses for use with rat
and mouse islet isolations.
Rodent HEPATOCYTE APPLICATIONS We
recommend: Defined Enriched Collagenases
DE Collagenase 60 or 600
DE Collagenase 10 or 100
These recommendations are dependent on
the protocol used in the isolation procedure.
(Recommended method: Seglen’s Two
-step Isolation Method 1,2.
To read more, please go to: https://
1,2: Seglen P.O. (1976) Preparation of isolated rat liver
cells. Methods in Cell Biology 13, 29-83.;
2: Seglen PO. Isolation of hepatocytes. In: Celis JE, ed. Cell
biology : a laboratory handbook. San Diego :: Academic
DNs-Rh is a novel fluorogenic substrate for GSTs (Glutathione S-Transferases). Compared to conventional reagents,
DNs-Rh can be used for live cell imaging with high specificity and sensitivity discovered by Dr. Hiroshi Abe, Nagoya
As many studies suggested, expression
level of GSTs is significantly
increased in cancer cells, GSTs are
considered as anti-cancer drugresistant
enzymes in malignant
cancer cells through the neutralization
of drugs. To understand biological
functions of GSTs, tools for
monitoring GST activity are very
important. Although several reagents
including CDNB (conventional
GSTs probe) for this purpose have
been developed, no tool to measure
intracellular GST activity was
DNs-Rh is a rhodamine 110 derivative which protected by DNs (dinitrobenzenesulfonamide).
This probe emits very low fluorescence (quantum yield = 0.0007) at normal state. After deprotected by thiols, rhodamine
110 is released and exhibits strong fluorescence (quantum yield = 0.645, S/N ratio ~900).
DNs-Rh can be used as a fluorogenic GST activity assay probe. An important advantage of this probe is high cellpermeability
and this probe is applied to intracellular GST activity under live cell condition.
As DNs group is a well characterized substrate for various types of GST members, DNs-Rh is able to monitor pan-
GST activity both in cell and in vitro.
These cells are endothelial-like cells in a sponge-like connective tissue located near the front
of the eye and are responsible for regualting eye pressure by controlling drainage of fluid into tubes that flow into
the bloodstream. Damage and dysfunction of TM have clinical significance. For instance, hypoxia increases DNA
methylation, accompanied by altered gene expression, whereas during the normal aging process, TM number decreases,
and senescent cells accumulate. In Glaucoma, a leading cause of irreversible blindness, decreased fluid
outflow causes an elevation of intraocular pressure and progressive loss of retinal ganglion cells. Physical changes
to the TM include increased fibrosis, fibronectin accumulation, and expression of ECM cross-linking enzymes.
Choose your Trabecular Meshwork Cells listed below or contact us for custom human or animal ocular cell types for
your research projects or applications: Ask Peter for more details email@example.com
Canine Trabecular Meshwork Cells: CnTMC, Human Trabecular Meshwork Cells: HnTMC, Porcine Trabecular
Meshwork Cells: PnTMC
More New Products
PELOBiotech‘s Premium-brand Cellovations®
launched NEW Media
Human Hepatocyte Medium - 2D (HHM-2D) and Human Hepatocyte
Medium - 2D (HHM-3D)
HHM-2D and HHM-3D are complete media formulated for optimal cultivation of all hepatocytes,
human and animal, etiher to grow in 2D after plating or to build spheroids (3D).
They are serum-free and growth factor containing culture media and contain essential and
non-essential amino acids, vitamins, organic and inorganic compounds, hormones, trace
minerals together with the growth factor containing supplements. They contain NO FCS. The
media are delivered as Kit with separate supplements. The media may be used for functionel
long-term cultivation of hepatocytes to study hepatotocxity of chemicals and drug-drug
interactions, or in hepatocyte bioreactors. Please be aware that hepatocytes do not proliferate.
They may be used to study hepatotocxity of chemicals and drug-drug interactions, or in hepatocyte
bioreactors. Please ask Lothar for further Details.
Cat. #: PB-MH-053-5990-2D or PB-MH-053-5990-3D
PELONews: Refer a friend
We are convinced
that we convince
you with our expertise
So if you are already
recommend us to
a colleague and
friend – you will
get a nice thankyou
reward. More details?
Call us now.
Ipita: Meet us at the
IPITA 2019 July, 02-
05 July, 2019 in Lyon,
France. You may
meet our CEO Dr.
Peter Frost at the
booth of our partner
Vitacyte at the
7 th World Congress of
PANCREAS & ISLET
Association. It´s a big
family meeting, so join
us now and meet Vitacytes‘s
McCarthy at booth#8.
Testimonials: All that matters is you!
We are so happy, that you are our customer. And even more if we
may use your words to be recommended to other customers and
scientists: Because we can say a lot about us —but all that matters
Enjoy your daily coffee or pot of tea with this great and unique
cup—made with love and only for you. Send us your favorite color
(and if you like a favorite short citation)and you will have this gift
asap on your desk. Thank you.
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comply to the new GDPR guidelines.
Dr. Peter Frost,
Dr. Lothar Steeb