Instructions for use

InviMag® Universal Kit/ KF96

Language: EN

7450300200 5 x 96 preparations Invitek Molecular GmbH

7450300250 Robert-Rössle-Straße 10
13125 Berlin
Germany
Important notes
Thank you for purchasing the InviMag® Universal Kit/KF96 from Invitek Diagnostics.
The product serves the purpose of semi-automated isolation of nucleic acids (genomic DNA,
bacterial DNA, viral DNA/RNA) from a variety of clinical samples using magnetic beads
technology.

WARNING! Improper handling and use for other than the intended purpose can cause danger
and damage. Therefore, we ask you to read through these instructions for use and follow them
carefully. Always keep them handy. To avoid personal injury, also observe the safety
instructions.

All versions of the instructions for use can be found on our website for download or can be
requested from us: www.invitek.com

Contact:
Technical support:
techsupport@invitek.com

GERMANY
Robert-Rössle-Str. 10, 13125 Berlin, Germany
+ 49 30 9489 2908

PORTUGAL
Zona Industrial de Tondela, ZIM II, Lote 6, 3460-070 Tondela, Portugal
+351 232 817 817

© 2023 Invitek Molecular, all rights reserved.

The kit is in compliance with REGULATION (EU) 2017/746 on in vitro diagnostic medical devices. But it is not for in-
vitro diagnostic use in countries where the REGULATION (EU) 2017/746 on in vitro diagnostic medical devices is not
recognized.

Trademarks: InviSorb®, PSP®, InviMag®, Eppendorf®. Registered marks, trademarks, etc. used in this document, even
when not specifically marked as such, are not to be considered unprotected by law.

InviGenius®, InviMag®, InviSorb®, Invitek®, InviTrap®, MSB®, PSP®, RTP® are registered trademarks of Invitek
Molecular GmbH.
Table of Contents
1. Safety instructions................................................................................................................ 3
2. Product information ................................................................................................................ 4
2.1 Kit contents ...................................................................................................................... 4
2.2 Reagents and equipment to be supplied by user ............................................................. 5
2.3 Storage, appearance and shelf life ................................................................................. 6
2.4 Intended use ...................................................................................................................... 6
2.5 Product information and specifications ............................................................................. 7
2.6 Principle and procedure .................................................................................................. 8
3. Nucleic acid extraction with the InviMag® Universal Kit KF/96 .......................................... 9
3.1 Before starting a protocol ................................................................................................ 9
3.2 Sampling and storage of starting material .................................................................... 10
3.3 Preparation of starting material ..................................................................................... 11
3.3.1 Serum, plasma, other cell-free body liquids ...................................................... 11
3.3.2 Blood................................................................................................................... 11
3.3.3 Swabs ................................................................................................................. 11
3.3.4 Stool samples (supernatant) .............................................................................. 12
3.3.5 Cultivated bacteria.............................................................................................. 12
3.3.6 Urine ................................................................................................................... 12
3.3.7 Tracheal secrete, BAL, sputum.......................................................................... 13
3.3.8 Cell culture supernatants ................................................................................... 13
3.4 Short protocol InviMag® Universal Kit/KF96 ................................................................. 14
3.5 Preparing and loading the KingFisher TM Flex .............................................................. 15
4. Appendix ........................................................................................................................... 16
4.1 Step-by-step protocol of the KingFisherTM Flex ............................................................ 16
4.2 Troubleshooting............................................................................................................. 19
4.3 Warranty ........................................................................................................................ 20
4.4 Symbols used on product and labeling ......................................................................... 20
4.5 Further documents and supplementary information ..................................................... 21
4.6 Ordering information ..................................................................................................... 21
 1. Safety instructions
Ensure that anyone using this product has received instructions in general safety practices for
laboratories and the safety information provided in this document.

• When and while working with chemicals, always wear protective clothing, disposable
gloves and safety glasses.
• Always change pipette tips between liquid transfers. To avoid cross-contamination, we
recommend the use of aerosol-barrier pipette tips.
• Do not reuse any consumables.
• Discard gloves if they become contaminated.
• Do not combine components of different kits unless the lot numbers are identical.
• Avoid microbial contamination of the kit reagents.
• To minimize the risk of infections from potentially infectious material, we recommend
working under laminar airflow until the samples are lysed.

Before handling chemicals read and understand all applicable safety data sheets (MSDS).
These are available online at www.invitek.com.
Dispose of kit residues and waste fluids in accordance with your country's regulations, again
refer to the MSDS. Invitek Molecular has not tested the liquid waste generated by the kit for
residual infectious materials. Contamination of the liquid waste with residual infectious
materials is highly unlikely but cannot be excluded completely. Therefore, liquid waste must
be considered infectious and must be handled and disposed of according to local safety
regulations.

European Community risk and safety phrases for the components of the InviMag® Universal
Kit/KF96 to which they apply are listed below as follows:

Proteinase K Lysis Buffer HLT

Danger Warning
H315-H319-H334-H335-P280-P305+P351+P338 H302-H315-H319-P280-P305+P351+P338

H302: Harmful if swallowed.

H315: Causes skin irritation.
H319: Causes serious eye irritation.
H334: May cause allergy or asthma symptoms or breathing difficulties if inhaled.
H335: May cause respiratory irritation.
P280: Wear protective gloves/protective clothing/eye protection/face protection.
P305+P351+P338: IF IN EYES: Rinse cautiously with water for several minutes. Remove contact
lenses, if present and easy to do. Continue rinsing

Emergency medical information can be obtained 24 hours a day from infotrac,

www.infotrac.net:

outside of USA: 1 – 352 – 323 – 3500

in USA: 1 – 800 – 535 – 50

InviMag® Universal Kit/KF96 3 EN-v2-2023

 2. Product information
2.1 Kit contents
InviMag® Universal Kit/ KF96
InviMag® Universal Kit/KF96
w/o plastic
5 x 96 preparations
5 x 96 preparations
Catalogue No. 7450300200 7450300250
Lysozyme Buffer 15 ml/bottle 15 ml/bottle
Lysozyme 150 mg/vial 150 mg/vial
Lysis Buffer HLT 120 ml/bottle 120 ml/bottle
Carrier RNA 10 x 1.2 ml working solution/vial 10 x 1.2 ml working solution/vial
RNAse Free Water 2 x 15 ml/bottle 2 x 15 ml/bottle
Proteinase K 10 x 1.1 ml working solution/vial 10 x 1.1 ml working solution/vial
SNAP Solution 10.5 ml/bottle 10.5 ml/bottle
Binding Solution empty bottle empty bottle
(fill with 99.7% Isopropanol) (final volume 120 ml) (final volume 120 ml)
360 ml/bottle 360 ml/bottle
Wash Buffer HLT
(final volume 600 ml) (final volume 600 ml)
180 ml/bottle 180 ml/bottle
Wash Buffer II
(final volume 600 ml) (final volume 600 ml)
150 ml/bottle 150 ml/bottle
Wash Buffer M
(final volume 600 ml) (final volume 600 ml)
Elution Buffer M 60 ml/bottle 60 ml/bottle
KF96 Tip Comb for DW
5 tip combs -
magnets
2.0 ml Deep Well Plate 5 x 4 plates -
200 µl Elution Plate 5 x 2 plates -
Sealing Foils 10 foils 10
1.5 ml Receiver Tubes 10 x 50 pieces 10 x 50 pieces
Short Protocol 1 leaflet 1 leaflet

InviMag® Universal Kit/KF96 4 EN-v2-2023

 2.2 Reagents and equipment to be supplied by user
Lab equipment:
• KingFisherTM Flex with 96 Deep-Well Head and consumables:
KingFisher 96, Magnetic Particle Processor, 100-240V, 50/60Hz with Deep well
Head (available at ThermoFisher Scientific)

KingFisher 96 tip comb

KingFisher 96 KF plate (200 µl)
DeepWell Platte 2 ml, KingFisher

• Microcentrifuge
• Thermo shaker (37°C - 95°C)
• Measuring cylinder (250 ml)
• Disposable gloves
• Pipette and pipette tips (filter tips are recommended)
• Vortex mixer
• Optional: Reaction tubes (1.5 ml, 15 ml, 50 ml)

Liquids and solvents:

• DNase/RNase free water or 1 x PBS to adjust sample volume
• 96 - 100 % ethanol (non-denatured)
• Isopropanol*
• Optional (for respiratory samples with high viscosity): saturated acetylcysteine (ACC)
solution (200 mg/ml)

*The InviMag® Universal Kit/ KF96 is validated with 2-Propanol; Rotipuran® >99.7%, p.a.,
ACS, ISO (Order no. 6752) from Carl Roth
* Possible suppliers for Isopropanol:

Carl Roth Applichem Sigma

2-Propanol 2-Propanol für die Molekularbiologie 2-Propanol
Rotipuran® >99.7%, p.a., ACS, ISO Order no. A3928 Order no. 59304-1L-F
Order no. 6752

InviMag® Universal Kit/KF96 5 EN-v2-2023

 2.3 Storage, appearance and shelf life
Shelf life: All buffers and kit components should be stored at room temperature and have a
shelf life as indicated on the outer kit package label.
After opening, individual components of the kit, as well as components prepared accordingly
before first use, have a shelf life of 3 months.
Before each use, make sure that all components are at room temperature. If there are
temperature-related precipitates in the solutions, dissolve them by carefully warming (up to 30°C).
Room temperature (RT) is defined as a range from 15-30°C.
Wash Buffer M and Wash Buffer II: after adding ethanol, it should be firmly closed and stored
at room temperature.
Wash Buffer HLT and Binding Solution: after adding isopropanol, they should be firmly closed
and stored at room temperature.
Carrier RNA: once dissolved in DNase/RNase free water Carrier RNA must be stored at
- 20°C.
Proteinase K: once dissolved in DNase/RNase free water Proteinase K can be stored at
2 -8 °C for up to two months. For longer storage keep at –20 °C, freeze-thaw once only.
Lysozyme: lyophilized Lysozyme must be stored at 2 - 8 °C. Dissolved Lysozyme (aliquoting is
recommended) must be stored at -20°C.

2.4 Intended use

The InviMag® Universal Kit/KF96 is for the semi-automated simultaneous isolation and
purification of genomic DNA, bacterial DNA and viral DNA/RNA using magnetic beads
technology.

The kit can be used for a variety of human sample types, such as fresh or frozen venous whole
blood anticoagulated with EDTA or citrate or the respective plasma preparations, serum, rinsed
liquid from swabs, pretreated sputum, BAL, tracheal secrete, cultivated bacteria, supernatant
from stool suspension, cerebrospinal fluid, cell culture supernatants, biopsy material/tissue,
urine, and other cell-free body fluids.

The InviMag® Universal Kit/KF96 is validated for the use on a KingFisherTM Flex (Thermo
Fisher Scientific) instrument. Ensure correct function and configuration of the instrument
according to the manufacturer’s instructions. Improper use of the instrument may result in lower
yields and can potentially harm the instrument.

The product is not intended to be used with heparinized blood samples. The product is
intended for use by professionals only, such as laboratory technicians, physicians and
biologists trained in molecular biological techniques and in vitro diagnostic procedures

InviMag® Universal Kit/KF96 6 EN-v2-2023

 2.5 Product information and specifications
Starting material Yield Quality Time

up to 200 µl: Depending on genomic DNA 40-60 min,

• Serum, plasma, other sample (storage from Blood: depending
cell-free body liquids, and source) A260 : A280 on protocol
urine 1.8 – 2.1 (incl. lysis)
• swabs (dry, stabilized) Whole blood: 3-6 Other sample
• fresh or frozen blood µg for a leukocyte types: depending
(EDTA / citrate stabilized, content of 3 x 106 on sample type,
but not heparinized) to target nucleic
• supernatant from stool 1 x 107 cells/ml acids
suspensions
• cultivated bacteria
• tracheal secrete, BAL,
sputum
• cell culture supernatant

Yield and quality of purified nucleic acids depend on the sample type, sample source, transport,
storage, age, the virus titer and for blood samples also on the leukocyte count.
For determination of yield please note that nucleic acids purified with this kit contain Carrier RNA
(5 μg per 200 μl sample), which account for most of the nucleic acids present in the eluate.
Especially viral nucleic acids from biological sample material are usually very low concentrated
and therefore almost impossible to be quantified photometrically. Quantitative RT-PCR is
recommended for yield determination.
The kit is validated for leukocyte counts of 3x106 - 1x107 cells/ml. Excessively high cell counts
may lead to undesirable effects on the purification process. It is therefore recommended to
consider sample input volume as a parameter during the implementation of your in vitro
diagnostic protocol. If required, samples may be pre-diluted with PBS or DNase/RNase free
water prior to the isolation and purification process.

Downstream Applications:
Yield and quality of isolated nucleic acids are in general suitable for plenty of molecular-
diagnostic applications such as PCR techniques, NGS, hybridization methods and HLA typing.
Downstream applications should be performed according to the respective manufacturers’
specifications.

InviMag® Universal Kit/KF96 7 EN-v2-2023

 2.6 Principle and procedure
The KingFisherTM Flex instrument uses magnetic rods to transport paramagnetic beads with
bound nucleic acids through the different extraction phases: lysis, binding, washing and
elution. The semi-automated purification process provides a reproducible method for the
recovery of highly pure nucleic acids.

1. Lyse samples
Samples are lysed at elevated temperatures. Lysis is performed in the presence of Lysis Buffer
HLT, Proteinase K and optionally lysozyme to break bacterial cell walls and to digest proteins.
The addition of Carrier RNA is required for the enhancement and stabilization of viral DNA/RNA
recovery and to purify very small amounts of viral nucleic acids.

2. Bind nucleic acids

After sample lysis the instrument will pause to adjust binding conditions by adding Binding
Solution to the lysate. Additionally SNAP Solution, containing silica coated magnetic beads, is
added. Nucleic acids bind specifically to the magnetic beads.

3. Wash to remove residual contaminations

Contaminants are efficiently washed away in three washing steps using Wash Buffer HLT, Wash
Buffer M and Wash Buffer II, while nucleic acids remain bound to magnetic beads.

4. Elute nucleic acids

Nucleic acids are released from magnetic beads and are eluted in 100 µl Elution Buffer M.

InviMag® Universal Kit/KF96 8 EN-v2-2023

 3. Nucleic acid extraction with the InviMag® Universal Kit KF/96
3.1 Before starting a protocol
When using the kit for the first time make sure all buffers are prepared as indicated:
Buffer preparations prior first use:
Binding Solution (empty bottle): Fill 120 ml 99.7% isopropanol (molecular biology grade)
into the bottle, always keep the bottle firmly closed.
Wash Buffer HLT: Add 240 ml of 99.7% isopropanol to the bottle. Mix thoroughly, always
keep the bottle firmly closed.
Wash Buffer M: Add 450 ml of 96 -100% ethanol to the bottle. Mix thoroughly, always keep the
bottle firmly closed.
Wash Buffer II: Add 420 ml of 96 -100% ethanol to the bottle. Mix thoroughly, always keep the
bottle firmly closed.
Carrier RNA: Resuspend each tube in 1.2 ml DNase/RNase free water. Mix thoroughly until
completely dissolved.
Proteinase K: Resuspend each tube in 1.1 ml DNase/RNase free water. Mix thoroughly until
completely dissolved.
Lysozyme: add the whole vial Lysozyme powder to the bottle of Lysozyme Buffer. Mix
thoroughly until completely dissolved. Make sure to prepare directly before first extraction, use
freshly prepared solution. Once dissolved, prepare aliquots and store at -20°C until further
use.

Master Mix
For easier handling, we recommend preparing a master mix consisting of Lysis Buffer HLT,
Proteinase K and, if required, Carrier RNA. When preparing the master mix, it is recommended
to prepare a volume that exceeds the total number of reactions by 5 %.
Always prepare the master mix fresh and shortly before use.
Isolating genomic DNA, bacterial DNA and viral DNA/RNA:
Per sample 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA are required.
Isolating genomic DNA:
Per sample 220 µl Lysis Buffer HLT and 20 µl Proteinase K are required. The use of Carrier
RNA is not required.

Extraction control
Refer to the manufacturer’s instructions to determine the optimal amount of extraction control
for specific downstream applications.
Low volumes of extraction control (DNA or RNA) must be combined with the provided Carrier
RNA in one mixture. The vials with Carrier RNA contain 1.2 ml stock solution. Add the
respective amount of extraction control nucleic acid to the Carrier RNA, if a high volume is
necessary (> 25% of the total Carrier RNA Volume), replace the appropriate amount of
DNase/RNase free water during dilution of the Carrier RNA.
Alternatively, extraction controls can be added after lysis, at the Binding step.

InviMag® Universal Kit/KF96 9 EN-v2-2023

 3.2 Sampling and storage of starting material
For reproducible and high yields, the correct sample storage is essential. Yields may vary
depending on factors such as health of the donor, sample age, sample type, transport and
storage.
Repeated freeze-thaw cycles of samples should be avoided to prevent nucleic acid
degradation. In general, best results are obtained using fresh samples. It is recommended to
consider technical guidance such as e.g. CEN/TS and ISO standards on the pre-examination
process for molecular diagnostics under IVDR as highlighted in G. Dagher, et al.
(https://doi.org/10.1016/j.nbt.2019.05.002).

Serum, plasma, other cell-free body liquids: Serum or plasma derived from venous whole
blood (treated with anticoagulants like EDTA or citrate, but not with heparin), synovial fluid
samples or other cell-free body fluids can be used for extraction. Whole blood should not be
vortexed as to avoid hemolysis. Allow serum tubes to sit for at least 30 min before
centrifugation. Follow blood collection system instructions for preparation of serum or plasma.
It is recommended to separate plasma/serum through centrifugation within 12 h. Supernatants
obtained using systems without gel separator should be transferred to fresh sample tubes. For
short-term storage, samples can be kept on ice for 1-2 hours. For up to 24 h samples can be
stored at -20°C. For long-term storage, freezing samples in aliquots at –80°C is recommended.
Repeated freeze-thaw cycles may negatively affect sample integrity and cause e.g.
denaturation/precipitation of proteins, potentially resulting in reduced yield, quality or viral
titers. In addition, cryoprecipitates formed during thaw-freeze cycles can cause problems. If
cryoprecipitate is visible, centrifuge at 6.800 x g for 3 min. The clear supernatant should be
used immediately.

Blood: Blood samples (stabilized with EDTA or citrate but not heparinized) can be stored at
room temperature for 2-3 hours. For short-term storage (up to 24 h) samples should be stored
at 2-8°C. For long-term storage, freezing samples at -20°C or -80°C is recommended.

Swabs:
Dry swabs: prepare the samples as described in the corresponding sample preparation
method. Store dry at 4-8°C.
Swabs in stabilization medium: the stabilization liquid can be handled as cell-free body fluid.
Please note that some stabilization agents may cause a reduced yield due to incompatibility
with chemistry used in the kit. Store according to the manufacturer’s requirements.

Stool samples: Samples contain DNases and RNases which can quickly cause DNA and
RNA degradation. Therefore, samples should be stored frozen at – 80°C.

Cultivated bacteria: After cultivation bacteria must be pelleted and frozen at -20°C or -80°C
for long-term storage. Resuspension is described in the corresponding sample preparation
method.

Urine: Depending on bacteria titer and application a starting volume of 15-50 ml urine is
recommended. Centrifuge the sample to pellet bacteria and remove the supernatant
completely (urea contaminations can inhibit PCR reactions). For some applications fresh urine
can be used directly. For long-term storage, freezing samples at -20°C or -80°C is
recommended.
Tracheal secrete, BAL, sputum: Samples contain DNases and RNases, which can quickly
cause DNA and RNA degradation. Therefore, samples should be stored frozen at – 80°C.
InviMag® Universal Kit/KF96 10 EN-v2-2023
Cell culture supernatants: Prepare supernatant samples like other cell-free body fluid
samples described in the corresponding sample preparation method. For long-term storage,
freezing samples at -20°C or -80°C is recommended.

3.3 Preparation of starting material

In the following the preparation of the sample lysis for different starting materials is described.
After loading the Lysis Plate with the respective sample and reagents, please proceed as
described in 3.5 “Preparing and loading the KingFisherTM Flex to continue with the extraction
procedure.

3.3.1 Serum, plasma, other cell-free body liquids

Always mix the sample well before extraction.
Transfer 200 µl sample into a cavity of the Lysis Plate. If the sample volume is below 200 µl,
adjust with 1 x PBS Buffer or DNase/RNase free water to a final volume of 200 µl.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA (optional for preparation
of genomic DNA) to each sample or add 240 µl Master Mix.
For the preparation of bacterial DNA, the addition of 20 µl Lysozyme is recommended.
Lysozyme needs to be added to the Lysis Plate before adding samples or other reagents.

3.3.2 Blood
Always mix the sample well before extraction.
Transfer 200 µl sample into a cavity of the Lysis Plate.
For extraction of genomic DNA add 220 µl Lysis Buffer HLT and 20 µl Proteinase K to each
sample or add 240 µl Master Mix.

3.3.3 Swabs
a) Dry Swabs
Rinse the swabs in a suitable vial in the lowest possible volume of PBS or DNase/RNase-free
water (for nasopharyngeal swabs about 400 µl, for oral swabs about 600 µl). Squeeze the
swab to the inner wall of the vial to obtain as much sample as possible.
Use 200 µl of the rinsed solution for extraction.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA (optional for preparation
of genomic DNA) to each sample or add 240 µl Master Mix.
Alternatively, swabs can be rinsed directly in 200 µl DNase/RNase free water plus Master Mix.
Insert swabs to single cavities of the lysis plate and incubate for 5-10 min at RT, mix
occasionally. Take care to avoid cross contamination.
For the preparation of bacterial DNA, the addition of 20 µl Lysozyme is recommended.
Lysozyme needs to be added to the Lysis Plate before adding samples or other reagents.

InviMag® Universal Kit/KF96 11 EN-v2-2023

 b) Swabs in stabilization medium
Use 200 µl of the stabilization solution for extraction.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA (optional for preparation
of genomic DNA) to each sample or add 240 µl Master Mix.
For the preparation of bacterial DNA, the addition of 20 µl Lysozyme is recommended.
Lysozyme needs to be added to the Lysis Plate before adding samples or other reagents.
Some stabilization media may interfere with the lysis reaction (if you have any questions,
please refer to the FAQ or contact support).

3.3.4 Stool samples (supernatant)

a) Extraction of nucleic acids from viruses
To prepare supernatant transfer 100 µl / 100 mg stool sample into a 2 ml vial and add 900 µl
DNase/RNase free water. Vortex for 30 s followed by a 1 min centrifugation step at 12.000 x
g.
Transfer 200 µl supernatant into a cavity of the Lysis Plate. Avoid solid particles in the sample.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA to each sample or add
240 µl Master Mix.
b) Extraction of bacterial DNA
To prepare supernatant transfer 100 µl / 100 mg stool sample into a 2 ml vial and add 300 µl
DNase/RNase free water. Vortex for 30 s followed by a 30 s centrifugation step at 1.000 x g.
Transfer 200 µl supernatant into a cavity of the Lysis Plate. Avoid solid particles in the sample.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA to each sample or add
240 µl Master Mix.
For the preparation of bacterial DNA, the addition of 20 µl Lysozyme is recommended.
Lysozyme needs to be added to the Lysis Plate before adding samples or other reagents.

3.3.5 Cultivated bacteria

Transfer 1ml of a bacterial overnight culture into a 2.0 ml Safe-Lock-Tube. Centrifuge for 2 min
at 10.000 x g and remove the supernatant completely. Resuspend the pellet in 200 µl PBS
Buffer and transfer the sample to the Lysis Plate.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA to each sample or add
240 µl Master Mix.
For the preparation of bacterial DNA, the addition of 20 µl Lysozyme is recommended.
Lysozyme needs to be added to the Lysis Plate before adding samples or other reagents.

3.3.6 Urine
Depending on bacteria titer and application a starting volume of 15-50 ml urine is
recommended. Centrifuge the sample to pellet bacteria and remove the supernatant
completely (urea contaminations can inhibit PCR reactions). Resuspend the bacteria pellet in
200 µl PBS Buffer.
For some applications 200 µl of fresh urine can be used directly.
Transfer the sample to the Lysis Plate and add 20 µl Lysozyme.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA to each sample or add
240 µl Master Mix.

InviMag® Universal Kit/KF96 12 EN-v2-2023

 3.3.7 Tracheal secrete, BAL, sputum
a) Non-viscous or low viscosity samples
Always mix the sample well before extraction.
Transfer 200 µl sample into a cavity of the Lysis Plate.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA (optional for preparation
of genomic DNA) to each sample or add 240 µl Master Mix.
For the preparation of bacterial DNA, the addition of 20 µl Lysozyme is recommended.
Lysozyme needs to be added to the Lysis Plate before adding samples or other reagents.
b) Isolation of bacterial DNA from viscous samples
Transfer 150 µl of the sputum sample or 1 ml tracheal secrete or BAL into a Safe- Lock-Tube
and add 150 µl or 1 ml saturated acetylcysteine (ACC) solution respectively (ratio sample to
buffer must be 1:1).
Incubate for 10 min at 95°C while continuously shaking.
Centrifuge at 10.000 x g for 5 min. Discard the supernatant.
Resuspend the bacterial pellet in 200 µl PBS or DNase/RNase free water and transfer the
sample to the Lysis Plate, add 20 µl Lysozyme.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA to each sample or add
240 µl Master Mix.
c) Isolation of viral DNA/RNA from viscous samples
Transfer 150 µl of the sample into a Safe-Lock-Tube and add 150 µl saturated acetylcysteine
(ACC) solution to the sample (ratio sample to buffer must be 1:1).
Incubate for 10 min at 95°C while continuously shaking.
Allow the sample to cool down.
Transfer 200 µl sample into a cavity of the Lysis Plate.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA to each sample or add
240 µl Master Mix.

3.3.8 Cell culture supernatants

Transfer 200 µl sample into a cavity of the Lysis Plate.
Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K and 20 µl Carrier RNA to each sample or add
240 µl Master Mix.
For the preparation of bacterial DNA, the addition of 20 µl Lysozyme is recommended.
Lysozyme needs to be added to the Lysis Plate before adding samples or other reagents.

InviMag® Universal Kit/KF96 13 EN-v2-2023

 3.4 Short protocol InviMag® Universal Kit/KF96

Refer to chapter 3.5 “Preparing and loading the KingFisherTM Flex“

Transfer 200 µl sample into a cavity of the Lysis Plate. Add 200 µl Lysis Buffer HLT, 20 µl Proteinase K
and 20 µl Carrier RNA (optional for genomic DNA). For the preparation of bacterial DNA, the addition of
20 µl Lysozyme is recommended. Lysozyme needs to be added to the Lysis Plate before adding
samples or other reagents.
Prepare all plates as described:
Lysis Plate: Refer to 3.3 “Preparation of starting material” for sample specific pre-treatment.
Washing Plate_1: Add 900 µl Wash Buffer HLT to a 2.0 ml Deep Well Plate
Washing Plate_2: Add 900 µl Wash Buffer M to a 2.0 ml Deep Well Plate
Washing Plate_3: Add 1000 µl Wash Buffer II to a 2.0 ml Deep Well Plate
Elution Plate: Add 100 µl Elution Buffer M to the Elution Plate (same size as Tip Plate)
Tip Plate: Insert the KF96 Tip Comb for DW magnets on a Tip Plate. Elution Plates and Tip
Plates are identical, use one Elution Plate as a Tip Plate.

The following steps are performed on the KingFisherTM instrument:

Lysis of the sample

After lysis, during the pause step,

add 230 µl Binding Solution and 20 µl SNAP Solution

Nucleic acids bind to magnetic particles

Washing of the particle fixed nucleic acids

Magnetic separation

Elution of nucleic acids

Magnetic Separation

Pure nucleic acids

InviMag® Universal Kit/KF96 14 EN-v2-2023

 3.5 Preparing and loading the KingFisher TM Flex
When operating the KingFisherTM Flex make sure you have read and understood the
manufacturer’s instructions.
1. Determine the number of needed reactions including controls and prepare all plates
needed for the purification procedure as follows. Label the short side of each plate
accordingly.
For handling extraction controls please refer to chapter 3.1 “Before starting a
protocol”.

Plate setup for KingFisher TM Flex:

Lysis Plate refer to chapter 3.3 “Preparation of starting material” for sample specific
pretreatment.
Washing plate_1 Add 900 µl Wash Buffer HLT into the cavities of a Deep Well Plate
Washing plate_2 Add 900 µl Wash Buffer M into the cavities of a Deep Well Plate
Washing plate_3 Add 1.000 µl Wash Buffer II into the cavities of a Deep Well Plate
Elution Plate Add 100 µl Elution Buffer M into the cavities of an Elution Plate

2. Lysis Plate: Prepare the Lysis Plate according to the sample type, as described in
“Preparation of starting material”.
3. Switch on the KingFisherTM Flex instrument
4. Tip Plate: Place the KF96 Tip Comb for DW magnets on the Elution Plate (Tip Plate).
5. Choose an assay for starting the run (all assays are available for download at the Invitek
Diagnostics web page).
InviMag_Universal_KF96_V2: protocol for extraction of genomic DNA and nucleic
acids of all kinds of pathogens including difficult to lyse pathogens.
InviMag_Universal_KF96_Virus: protocol for the extraction of nucleic acids from
viruses from plasma or swab samples.
6. Load prepared plates into the position specified in the display of the instrument
7. “START” the run
8. After the lysis steps, the instrument pauses and 230 µl Binding Solution and 20 µl
SNAP Solution must be added to each reaction.
Note: Mix SNAP Solution before use by vortexing vigorously!
9. Place the plate back to the instrument (watch out for correct plate orientation) and
continue the run by pressing the “START” button. The instrument will now continue
the purification process without any further user interaction.
10. After extraction: A transfer of purified nucleic acids to 1,5 ml Receiver Tubes
(provided) is recommended.
Keep nucleic acids at -20°C or -80°C until further use.

InviMag® Universal Kit/KF96 15 EN-v2-2023

 4. Appendix
4.1 Step-by-step protocol of the KingFisherTM Flex

Protocol: InviMag_Universal_KF96_V2

InviMag® Universal Kit/KF96 16 EN-v2-2023

InviMag® Universal Kit/KF96 17 EN-v2-2023
InviMag® Universal Kit/KF96 18 EN-v2-2023
 4.2 Troubleshooting

Problem Possible cause Recommendation

Low amount Insufficient cell lysis Increase lysis time in the provided run file.
of nucleic Reduce amount of starting material.
acids
Incomplete elution Increase the volume of Elution Buffer M.
change the modified volume in the provided run file, too.

Low amount of SNAP Mix SNAP Solution thoroughly before use.

Solution

Low nucleic acid- Elute with in a lower volume (minimum 50 µl) of Elution
concentration in the Buffer M. Change the modified volume in the run file, too.
sample

Incorrect storage of starting Ensure that starting material is appropriately stored.

material Avoid repeated thaw-freeze cycles of the sample material.

Wash Buffers were Ensure, that the correct amount of ethanol/isopropanol is

incorrectly prepared added to the Wash Buffers and that all solutions are
stored firmly closed.

Degraded Incorrect storage of starting Ensure that the starting material is appropriately stored.
nucleic acids material Avoid repeated thaw-freeze cycles of the sample material.

Ensure that the starting material is stored at appropriate

Old material
conditions (–20°C/-80°C).

Nucleic acids Ethanol carryover during Increase time of drying step for removal of ethanol in the
do not elution run file.
perform well
in Salt carry-over during Check the Wash Buffers for salt precipitates. If there are
downstream- elution any precipitates visible, solve them by carefully warming
applications up to 30°C
(e.g. real-time Ensure that the Wash Buffers are at room temperature
PCR or NGS) before use.

No PCR result for genomic Due to the very gentle isolation procedure, it may happen
DNA that isolated genomic DNA forms a cluster. To avoid this,
the primary PCR denaturation step at 95°C should be
prolonged to 5 min

Magnetic Residues of magnetic Centrifuge eluted nucleic acids at full speed for 1 min and
beads carry- particles in eluted transfer supernatant to a new tube.
over extraction

InviMag® Universal Kit/KF96 19 EN-v2-2023

 4.3 Warranty
Invitek Molecular guarantees the correct function of the kit for applications described in this
manual and in accordance with the intended use. In accordance with Invitek Molecular’s EN ISO
13485 certified Quality Management System the performance of all kit components has been
tested to ensure product quality.
Any problems, incidents or defects shall be reported to Invitek Molecular immediately upon
detection. Immediately upon receipt, inspect the product to ensure that it is complete and intact.
In the event of any discrepancies, you must inform Invitek Molecular immediately in writing.
Modifications of the kit and protocols and use that deviate from the intended purpose are not
covered by any warranty.
Invitek Molecular reserves the right to change, alter, or modify any product to enhance its
performance and design at any time.
Invitek Molecular warrants products as set forth in the General Terms and Conditions available
at www.invitek.com. If you have any questions, please contact techsupport@invitek.com.

4.4 Symbols used on product and labeling

Manufacturer
Lot number

Unique identifier of a medical device

Catalogue number
Expiry date
Consult operating instructions
Temperature limitation
Do not reuse
Amount of sample preparations
in vitro diagnostic medical device

InviMag® Universal Kit/KF96 20 EN-v2-2023

 4.5 Further documents and supplementary information
Visit www.invitek.com for further information on:

• FAQs and troubleshooting tips

• Manuals in different languages
• Safety data Sheets (MSDS)
• Web support
• Product videos
• Run-Files for KingFisher™ Flex and KingFisher TM 96

If, despite careful study of the operating instructions and further information, you still require
assistance, please contact us at techsupport@invitek.com or the dealer responsible for you.

4.6 Ordering information

Product Package Size Catalogue No.

InviMag® Universal Kit /KF96 5 x 96 preparations 7450300200
InviMag® Universal Kit /KF96 w/o plastic 5 x 96 preparations 7450300250

Revision history

Revision Date Description

EN-v1-2022 2022-05-18 New document

EN-v2-2023 2023-04-17 Update contact details and

corporate design to reflect
the company's rebranding.

InviMag® Universal Kit/KF96 21 EN-v2-2023

 PORTUGAL
Zona Industrial de Tondela, ZIM II, Lote 2 e 6
63460-070 Tondela
Portugal

Phone: +351 232 817 817

GERMANY
Robert-Rössle-Str. 10
13125 Berlin
Germany

Phone: +49 30 9489 2908

info@invitek.com
www.invitek.com

2023-04-17 EN-v2-2023