Application Note 4Guidelines for recovering cells andspheroids from GrowDex®
using GrowDase™ enzyme
GROWDASE IS USED FOR THE
ENZYMATIC DEGRADATION OF
GROWDEX

GrowDase™ is a purified mixture of cellulase enzymes that have been specifically prepared
to degrade the nanocellulose fibrils present in GrowDex®, reducing them to soluble
glucose. This simple enzymatic removal procedure allows cells to be recovered from thematrix efficiently, without adverse effects and retain any 3D structure such as a spheroid ororganoid fully intact. The degradation process is simple; enzyme is mixed with cell culture
media and incubated with GrowDex at 37°C until the hydrogel has been dissolved away(Fig. 1).
The amount of GrowDase required for cell recovery is dependent on the amount ofGrowDex (cellulose) present in the experimental well. It is recommended that GrowDase
is used at a working concentration of 300 µg/mg (µg enzyme/mg cellulose). An equalvolume of working concentration GrowDase to GrowDex/cell matrix volume present in theexperimental well should be used, i.e. if 100 µL of GrowDex/cell matrix is present in theexperimental well then 100 µl of GrowDase should be added to that well.

Figure 1. Schematic presentation ofGrowDex degradation with GrowDase

PROCEDURE FOR REMOVINGGROWDEX BY ENZYMATIC
DEGRADATION

1.

Calculate the amount of cellulose present in the sample well using the followingequation. NOTE: 100 µL of 1% GrowDex contains 1 mg cellulose.

2.

Prepare the working concentration of GrowDase by diluting the stock solution withculture media.
Pipette the diluted GrowDase onto the top of the sample in the microplate.
Incubate the plate at 37°C until the hydrogel has fully degraded.
Recover the cells from the well using standard techniques.

3.
4.
5.

EXAMPLE EXPERIMENTAL
PROCEDURE

DILUTION TABLESAMPLE: 80 µL of 0.9% GrowDex/cell mix per well in a 96-well microplate
1.

Amount of GrowDex present in the sample:
80 µL x 0.9 / 100 = 0.72 mg GrowDex/ sample well.

2.

Amount of GrowDase needed to degrade the cellulose in the sample :0.72 mg x 300 µg/mg = 216 µg GrowDase enzyme.

3.

Volume of GrowDase stock solution (10 mg/mL) needed:
216 µg / 10 µg/µL = 21.6 µL GrowDase enzyme stock solution.

4.

Prepare the working concentration of GrowDase by diluting the stock solution withculture media:
80 µL - 21.6 µL = 58.4 µL culture media for dilution.

5.

Pipette the diluted GrowDase onto the top of the sample, incubate at 37°C until thehydrogel has fully degraded and recover cells from the well using standard techniques.

Volumes of GrowDase enzyme stock solution (10 mg/mL) and cell culture medium required
for the preparation of 100 µL of 300 µg/mg enzyme working solution for the degradation of
100 µL GrowDex in different concentrations from 0.3% to 1%.

GROWDEX CONC IN
100 µL OF SAMPLEAMOUNT OF
GROWDASE
NEEDEDVOLUME OFGROWDASE
STOCK SOLUTION(10 MG/ML)

VOLUME OF
CULTURE MEDIA

1%300 µl

30 µl

70 µl

0.9%

270 µl

27 µl

73 µl

0.8%

240 µl

24 µl

76 µl

0.7%

210 µl

21 µl

79 µl

0.6%

180 µl

18 µl

82 µl

0.5%

150 µl

15 µl

85 µl

0.4%

120 µl

12 µl

88 µl

0.3%

90 µl

9 µl

91 µl

ORDERING INFORMATIONCATALOGUE CODEDESCRIPTIONQUANTITY (ml)100 103 005GrowDex® (supplied in syringe)

5.0

100 103 010

GrowDex® (supplied in syringe)

10.0

100 103 305

GrowDex® multipack (supplied in syringe)

3 x 5.0

900 102 002

GrowDase Enzyme

2.5™

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Or contact us at biomedicals.sales@upm.com for a quotation or to place an order.

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