Preparing the Positive Product Control (PPC) Spike for

Pharma&Biotech
Preparing the Positive Product Control (PPC)
Spike for the QCL-1000™ Assay
A spiked aliquot of the test sample (or dilution) may be prepared
as follows:
Technical Tips
by Lonza Scientific Support
Product inhibition occurs when substances in the test sample interfere
with the Limulus Amebocyte Lysate (LAL) reaction. In the chromogenic
endpoint assay, this inhibition results in a lower final delta (∆) absorbance, indicating lower levels of endotoxin than what may actually be
present in the test sample. The absence of product inhibition should be
determined for each specific sample, either undiluted or corrected with
an appropriate dilution.
To verify the lack of product inhibition, an aliquot of test sample (or a
dilution of test sample) is spiked with a known amount of endotoxin
(e.g. 0.4 EU/ml). The spiked solution is assayed along with the un-spiked
samples and their respective endotoxin concentrations are determined.
The difference between the two calculated endotoxin values should
equal the known concentration of the spike ±25%.
Preparation of a 2.0 EU/ml stock to be used for spiking
X = The stated potency of the control standard endotoxin (CSE)
as stated on the kit Certificate of Analysis (CoA)
0.200 = Volume of CSE required
(X–2) / 10 = Volume of LAL Reagent Water (LRW) required
Example
X = 29
(29–2) / 10 = 2.7 ml LRW
2.7 ml LRW + 0.200 ml CSE = 2.9 ml stock at 2.0 EU/ml
Plate method #1
Transfer 10 µl of the 2.0 EU/ml solution into each of the Positive Product
Control (PPC) wells in the plate. To these wells add 50 µl of test sample (or
dilution). Each well will now contain 0.02 EU/well or 0.4 EU/ml. Mix gently.
Plate method #2
Place 50 µl of test sample (or dilution) into the PPC wells in the plate.
To these wells, add 10 µl of the 2.0 EU/ml solution. Each well will now
contain 0.02 EU/well or 0.4 EU/ml. Mix gently.
To calculate PPC recovery percent, subtract the average result of the unspiked wells from the average result of the spiked wells. Divide the difference by the value of the spike. In the example above, this spike value
would be 0.4 EU/ml. Multiply this result by 100.
(Spiked – Un-spiked)
× 100 = PPC % recovery (specification: 75 –125 %)
Spike value
If the test sample (or dilution) is found to be inhibitory to the QCL-1000™
Assay reaction, the sample may require further dilution or other pretreatment until the inhibition is overcome.
www.lonza.com
www.lonza.com/qcl1000
Contact Information
North America
Customer Service: +1 800 638 8174 (toll free)
[email protected]
Scientific Support: +1 800 521 0390 (toll free)
[email protected]
Europe
Customer Service: +32 87 321 611
[email protected]
Scientific Support: +32 87 321 611
[email protected]
International
Contact your local Lonza distributor
Customer Service: +1 301 898 7025
Fax: +1 301 845 8291
[email protected]
International Offices
Australia
+61 3 9550 0883
Belgium
+32 87 321 611
Brazil
+55 11 2069 8800
France0800 91 19 81 (toll free)
Germany0800 182 52 87 (toll free)
India
+91 40 4123 4000
Japan
+81 3 6264 0660
Luxemburg
+32 87 321 611
Singapore
+65 6521 4379
The Netherlands0800 022 4525 (toll free)
United Kingdom0808 234 97 88 (toll free)
Lonza Cologne GmbH – 50829 Cologne Germany
Unless otherwise noted, all trademarks herein are marks of the
Lonza Group Ltd or its affiliates. The information contained herein
is believed to be correct and corresponds to the latest state of
scientific and technical knowledge. However, no warranty is made,
either expressed or implied, regarding its accuracy or the results
to be obtained from the use of such information and no warranty
is expressed or implied concerning the use of these products. The
buyer assumes all risks of use and/or handling. Any user must
make his own determination and satisfy himself that the products
supplied by Lonza Group Ltd or its affiliates and the information
and recommendations given by Lonza Group Ltd or its affiliates
are (i) suitable for intended process or purpose, (ii) in compliance
with environmental, health and safety regulations, and (iii) will not
infringe any third party’s intellectual property rights.
© Copyright 2014, Lonza Cologne GmbH. All rights reserved.
RT-DS017 06/14

Download Report