ᲰᲨ‫୺ࠢف‬ዴȑǿȸȳƂࡑూɥƷදॖƃFor research use onlyᲬᲨPrimer design
 Appropriate primer design is essential for amplification by LAMP. For
the primer design, exclusive primer designing software, PrimerExplorer isavailable from Eiken GENOME SITE (URL;http://loopamp.eiken.co.jp/e/).
When use the highly purified primers, a rapid gene amplification canbe performed and stable reproducibility of the amplification can beobtained. The first screening for appropriate LAMP primers might notnecessarily require highly purified primers. However, after the primers
are determined, it is recommended to use purified FIP and BIP through
HPLC or better purification.

ᲫᲨӒࣖࢸƷȁȥȸȖƸᔟǛ᧏ƚƣƴŴ໲ҲϼྸӍƸ݅᧍ưƖǔȓȋȸȫƷᘥǛʚ᣻ƴ଀®

ƠᢘЏƴϼྸƠƯƘƩƞƍŵ‫ࠢف‬ငཋƷ᫠૝᧸ഥƷƨǊŴᑄ᫈ߩ㓙ߦࠝ࡯࠻ࠢ࡟࡯

ࡉಣℂߪⴕࠊߥ޿ߢߊߛߐ޿‫ޕ‬ᲬᲨᚾᕤȁȥȸȖƸ᳊᳊ŴǭȃȈǱȸǹƸኡǛɼƳ஬ឋƱƠƯƍǇƢŵ
ᲭᲨ̅ဇࢸƷஜԼǍܾ֥ӏƼ֥φ᫏ƸŴࡑూཋƷϼྸӏƼฌੈƴ᧙ƢǔඥࢷǍ൦ឋ൲ບ᧸DNA Amplification KitഥඥሁƴࢼƍŴӲ଀ᚨƷᝧ˓ƴƓƍƯϼྸƠƯƘƩƞƍŵ

This kit is for research use only. Do not use fordiagnosis or its assistance for clinical purpose.
Read this instruction carefully before use.

Ƃᝪᔺ૾ඥȷஊј஖᧓ȷѼᘺҥˮȷᙌԼǳȸȉƃᙌ Լ Ӹᝪᔺ૾ඥ

Loopamp DNA ‫ࠢف‬ᚾᕤǭȃȈ

Ყ20 é

ஊј஖᧓

1 ࠰᧓

Ჶย‫ܭ‬ƴƋƨƬƯƷදॖᲸ
ᲫᲨLAMP ӒࣖƸ᩼ࠝƴᥘ૓ƳӒࣖưƋǓŴ೅ႎᢡˡ‫܇‬Ǎ‫ࠢف‬ငཋƕಊࣇ᣽ưǋฆλƢǔƱᛚƬƨኽௐǛǋƨǒƢҾ‫׆‬ƱƳǔƓƦǕƕƋǓǇƢŵƜƷǑƏƳǳȳǿȟȍȸǷȧ

Ѽᘺҥˮ

ᙌԼǳȸȉ

48 ȆǹȈЎ

LMP20496 ȆǹȈЎ

LMP205

192 ȆǹȈЎ

LMP206

Ƃɼᙲ૨ྂƃ

ȳǛ‫ׅ‬ᢤƢǔƨǊƴŴ౨˳੔ӕƸஜᙌԼǛ̅ဇƢǔᢿ‫ދ‬ƱƸКƷᢿ‫ދ‬ưܱ଀ƢǔƔŴ1ᲣNotomi T. et al., : Nucleic Acids Research 28, No.12, e 63Ტ2000ᲣƋǔƍƸ౨௹ғ؏ǛЎлƠƯКƷǨȪǢưᘍƬƯƘƩƞƍŵ࣏ᙲƴࣖơƯŴǯȪȸȳ

2ᲣNagamine K. et al., : Clin. Chem. 47, No.9, 1742Ყ1743Ტ2001Უ

șȳȁƷ̅ဇӍƸ৖ᘥǍǢǤǽȬȸǷȧȳǬǦȳƷბဇሁưǳȳǿȟȍȸǷȧȳǛ

3ᲣMori Y. et al., : Biochem. Biophys. Res. Commun. 289, No.1, 150Ყ154Ტ2001Უ
4ᲣTomita N.et al., : Nat Protoc. 3, No.5, 877Ყ882᧸ƙਜ਼ፗǛƱƬƯƘƩƞƍŵ

5ᲣNagamine K. et al., : Molecular and Cellular Probes 16, No.3, 223Ყ229Ტ2002ᲣᲬᲨஜᙌԼǛӕǓৢƏᨥƴƸŴࣇဃཋǍఋᣠЎᚐᣞእᲢDNaseᲦRNaseᲣƷǳȳǿȟȍȸ

6Უଐஜኬᓏ‫˟ܖ‬ȐǤǪǻǤȕȆǣȸ‫ۀ‬Ճ˟Ჴଐஜኬᓏ‫ܖ‬ᩃᛏ, 54, No.3, 667Ყ715Ტ1999Უ

ǷȧȳǛᢤƚƯƘƩƞƍŵ
ᲭᲨBst DNA Polymerase Ƹ‫෇ڂ‬ƢǔƓƦǕƕƋǓǇƢƷưŴȞǹǿȸȟȃǯǹᛦᙌƷᨥƴƸນƠƘટਉƠƳƍưƘƩƞƍŵȜȫȆȃǯǹȟǭǵȸưટਉǛᘍƏ‫ئ‬ӳƸŴ1 ᅺᲭᲨReagents preparationƂCharacteristicsƃ
LAMP Ტ Loop Ყ mediated Isothermal Amplification Უ method is a gene
amplification method capturing the following characteristics : Ĭ The
amplification reaction proceeds under isothermal condition1 Უ, 2 Უ.,ĭ It has
extremely high specificity because of the use of 4 primers recognizing 6
distinct regions on the target.,Į It has high amplification efficiency andenables amplification within a shorter time., and į it produces tremendous
amount of amplified products which makes simple detection possible3Უ, 4Უ.
  This kit is for amplifying and detecting target gene sequence by the
LAMP method in combination with personally designed primers for LAMP, orprimer sets sold separately.

Ƃ Contents of the kitƃ     1Შ 2 × Reaction Mix.ᲢRMᲣè1௿ᄂ҄‫ܖ‬ఇࡸ˟ᅈ ƓܲಮႻᛩᆸӝ3Შ Distilled WaterᲢDWᲣè1ȕȪȸȀǤȤȫ㩷 㩷 㩷 0120Ყ308Ყ421

ƞƍŵŹ Sample reaction
48 tests 96 tests 192 tests

0.6 mLg1   g2    g4

4Შ Primer Mix. DNA è 2ᲢPM DNAᲣè1ᲮᲨȞǹǿȸȟȃǯǹƱǵȳȗȫ๋෩ǛฆԧࢸŴӒࣖ෩ƴൢණƕസƬƯƍǔƱບࡇย‫ܭ‬Ʒ

5Შ Positive Control DNA è 2ᲢPC DNAᲣè1

60L

g1   g2    g4

1.0 mLg1   g2    g2

25Lg1  Ყ   Ყ

0.1 mLg1  Ყ   ᲧᲶReagentsᲸ

Distilled WaterᲢDWᲣ
2gReaction MixᲢRMᲣ      PrimerᲴ FIP
BIP
LFè4
LBè4
F3
B3
Bst DNA Polymerase2Შ Bst DNA PolymeraseᲢBst DNA PolymeraseᲣè1ƂբƍӳǘƤέƃ

᧓g3 ‫ׅ‬Ǜӈ‫ܣ‬ƠƯƘƩƞƍŵ‫ݵ‬ŴᛦᙌƠƨȞǹǿȸȟȃǯǹƸƢƙƴ̅ဇƠƯƘƩ  

1Უ Take out the reagents stored at Ყ 20 é , and thaw them at room
temperature. Once the reagents are thawed, keep them on ice. After
thawed, tap the reagent tubes gently -- mixing (tapping) -- or invert thetube, or by vortex mixer at about 3 times for 1 second. After mixing well,centrifuge the tubes for a few seconds.
2Უ Preparation of master mix.ᲢOperate on iceᲣ.
The following amount of the component is required for one reaction.
When used in combination with Loopamp Primer Set, follow the
preparation instructions of each primer set.
ᲶAmountᲸ   
X
12.5
40
40
20
20
5
5
1.0

Total

L (ad q.s.)
L
pmol
pmol
pmol
pmol
pmol
pmol
L

23.0

LᲩtestૅᨦƱƳǓᛚЙ‫ܭ‬ƷҾ‫׆‬ƱƳǓǇƢƷưŴൢණƕဃơƳƍǑƏදॖƠƯƘƩƞƍŵൢè1ᲴThe notation on each reagent tube is shown inᲢණƕസƬƯƍǔ‫ئ‬ӳƴƸŴǹȔȳȀǦȳƠƯൢණǛӕǓᨊƍƯƘƩƞƍŵ

Უ.

è4ᲴLoop primers are not necessarily required. However, the use ofprimers shortens the amplification time by about one-third5Უ.

è2ᲴPrimer Mix. DNA and Positive Control DNA are not included in the 96ᲯᲨӒࣖࢸƷȁȥȸȖƷᔟƸൿƠƯ᧏ƚƳƍưƘƩƞƍŵཎƴӒࣖࢸƷȁȥȸȖǛᘺፗƔ

tests and 192 tests package.

ǒӕǓЈƢƱƖƴȁȥȸȖƷᔟƕ᧏ƔƳƍǑƏŴ॰᣻ƴӕǓЈƠƯƘƩƞƍŵ‫ࠢف‬ငŹ Control reactionƂPrincipleƃ

ཋƴǑǔǳȳǿȟȍȸǷȧȳƸᛚЙ‫ܭ‬ƷҾ‫׆‬ƱƳǔƹƔǓưƳƘŴย‫ܭ‬࿢‫ؾ‬ƦƷǋƷ

LAMP method is an isothermal nucleic acid amplification method using
4 kinds of primers and DNA polymerase with strand displacement activity.
Among 4 kinds of primers, two of them are inner primers, whose 3’ end and
5’ end are respectively designed to be complementary to two different
regions of the target sequence. When the primerᲧlinked strand is replicated,
resulting 3Þend structure to selfᲧanneals to the complementary region in selfᲧ
structure, it forms the loop structure at the end.
This stemᲧloop structure allow the 3Þ end to initiate selfᲧelongation andanother inner primer to anneal to its loop region to synthesize new DNA
strand with strandᲧdisplacement manner. Through the repetition of theseprocesses, the amplification proceeds, and this enables amplification tocontinue under isothermal condition with only one kind of enzyme.
For further details of the LAMP method, refer to Eiken GENOME SITEᲢURLᲵhttp://loopamp.eiken.co.jp/e/Უ.

Ǜ൲௨ƠŴ൲௨Ǜ‫ܦ‬μƴᨊӊƠƳƍᨂǓŴˌࢸദƠƍኽௐƕࢽǒǕƳƘƳǔӧᏡࣱƕƋǓǇƢŵᲰᲨᩓൢපѣሁưƷ‫ࠢف‬ငཋƷӕৢƍƸᢤƚƯƘƩƞƍŵ

ƂӕৢƍɥᲢүᨖ᧸ഥᲣƷදॖƃᲫᲨஜᙌԼƸŴ˳‫ٳ‬ᚮૺဇҔᕤԼưƸƋǓǇƤǜŵ
ᲬᲨ౨˳Ƹज़௨ƷүᨖƕƋǔǋƷƱƠƯදॖƠƯӕǓৢƍŴ࣏ᙲƳȐǤǪȏǶȸȉ‫ݣ‬ሊǛܱ଀ƠƯƘƩƞƍ 6ᲣŵᲭᲨᖩήႸᙻЙ‫଺ܭ‬ƴከ‫ٳ‬ዴༀ‫ݧ‬ᘺፗǛ̅ဇƢǔ‫ئ‬ӳŴȩȳȗǑǓ્‫ݧ‬ƞǕǔከ‫ٳ‬ዴᲢൈᓏዴᲣƸஊܹƳƷưŴከ‫ٳ‬ዴǛႺ੗ႸƴλǕǔƜƱƸᢤƚƯƘƩƞƍŵǇƨໜ໊ɶƷȩȳȗǛᙸǔ࣏ᙲƕƋǔƱƖƸŴ࣏ƣǬȩǹெǛᡫƢƔŴ࠼ࠢƷაᦟӍƸ᧸ᜱ᩿ǛƔƚƯЙ‫ܭ‬ƠƯƘƩƞƍŵ

ƂHow to useƃƂ̅ဇɥƷදॖƃ

ᲫᲨMaterials required but not provideᲫᲨஜᙌԼƸ࣏ƣᲧ20éư̬‫܍‬ƠƯƘƩƞƍŵᚾᕤƷэ҄Ǜ᧸ഥƢǔƨǊƴŴ̅ဇ଺Ƹ࣏

1ᲣSterilized tubes for master mix. preparationᲢ0.5 mL or 1.5 mLᲣ
2ᲣMicropipettes
3ᲣPipette tips with filterᲢDNase, RNase freeᲣ4ᲣLoopamp Reaction TubeᲢsold separatelyᲣ5ᲣAluminum rack for cooling tubes
6ᲣIceᲢcrushed iceᲣand ice box
7ᲣCentrifuge for microtube
8ᲣCentrifuge for 8Ყstrip tubes
9ᲣVortex mixerᙲƳᚾᕤƩƚǛረƔǒӕǓЈƠƯƝ̅ဇƘƩƞƍŵ
ᲢϵኽᗡᚐǛ 20 ‫ׅ‬ጮǓᡉƠƨኽௐưƸŴᚾᕤƷэ҄ƸǄƱǜƲᛐǊǒǕƯƓǓǇƤǜƕŴ໯ဇƳϵኽᗡᚐƸԼឋዜਤƷƨǊƴᢤƚƯƘƩƞƍŵᲣᲬᲨӒࣖȁȥȸȖƸ࣏ƣ‫ݦ‬ဇƷ Loopamp ӒࣖȁȥȸȖǛƝ̅ဇƘƩƞƍŵਦ‫ٳˌܭ‬ƷӒࣖȁȥȸȖǛ̅ဇƠƨ‫ئ‬ӳŴήᡢᢅࣱƷᢌƍƴǑǓᛚЙ‫ܭ‬ǛਔƘӧᏡࣱƕƋǓǇƢŵ
ᲭᲨӒࣖȁȥȸȖƸᄊ੷ƠǍƢƍƷưŴӕৢƍƴƸදॖƠƯƘƩƞƍŵ
ᲮᲨӒࣖȁȥȸȖƸဇƍǔЭƴǭǺȷȒȓሁƕ໯ƍƜƱǛႸᙻưᄩᛐƠƯƘƩƞƍŵӒࣖ

᲻ᲨFor RealᲧtime turbidity detection
Loopamp Realtime Turbidimeter è3ȁȥȸȖƴǭǺȷȒȓሁƕƋǔƱദƠƘย‫ܭ‬ưƖƳƍƹƔǓƔŴȁȥȸȖƷᄊ੷ƴǑǓᘺፗǛ൲௨ƢǔӧᏡࣱƕƋǓǇƢŵȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗӍƸǤȳǭȥșȸǿȸưŴஜᙌԼƷ̅ဇƴƋƨƬƯƸᢡˡ‫܇‬౨௹ƷჷᜤŴኺ᬴ǛஊƠƨ২ᘐᎍƷਦ‫ݰ‬ƷǋƱ

᲼ᲨFor Visual fluorescence detection
1ᲣLoopamp Fluorescent Detection ReagentᲢsold separatelyᲣ2ᲣLoopamp Realtime Turbidimeter è3, or incubator with hot bonnet
Ტtemperature accuracy withind0.5éᲣ3ᲣHeat blockᲢfor enzyme inactivationᲣè3
4ᲣUV lampᲢwavelength at 240᳸260nm or 350᳸370nmᲣè3
5ᲣProtective goggle or glass boardư౨௹Ǜܱ଀ƠƯƘƩƞƍŵ

è3ᲴFor the information about applicable instrument, reaction terminationƷӒࣖȖȭȃǯϋưȁȥȸȖƕᄊ੷Ơƨ‫ئ‬ӳƸŴӒࣖ෩ƕᘺፗஜ˳ǁ๨ЈƠŴᨊӊɧᏡƳ൲௨Ǎ૏ᨦƷҾ‫׆‬ƱƳǓǇƢŵ
ᲯᲨӒࣖȁȥȸȖŴȞǹǿȸȟȃǯǹᛦᙌဇ๒ᓏȁȥȸȖƴƸከ‫ٳ‬ዴༀ‫ݧ‬ƠƳƍưƘƩƞ
ƍŵከ‫ٳ‬ዴༀ‫ݧ‬ƴǑǔ‫٭‬ᑥȷ‫٭‬ឋሁưᛚƬƨኽௐǛǋƨǒƢ‫ئ‬ӳƕƋǓǇƢŵ
ᲰᲨᢡˡ‫܇‬౨௹ƷჷᜤǍኺ᬴ǛǋƨƳƍ‫ئ‬ӳŴ౨௹ኽௐƷЙ‫ܭ‬ǛᛚǔүᨖࣱƕƋǓǇƢƷ

function and condition for UV irradiation, refer to Eiken GENOME SITEᲢURL;http://loopamp.eiken.co.jp/e/Უ.

ᲱᲨஜᙌԼƷࣱᏡƴဌஹƠƳƍʙဌᲢદ˺૾ඥǛᛚƬƨ‫ئ‬ӳሁᲣƴǑǔᛚƬƨЙ‫ܭ‬ŴǇƨƦƷЙ‫ܭ‬ƴဌஹƠƯႆဃƠƨʙ᪮ƴ‫ݣ‬ƠƯŴࡴᅈƸɟЏƷᝧ˓Ǜ᝟ƍǇƤǜŵ
ᲲᲨஜᙌԼƸ̅ဇ஖ᨂϋƴ̅ဇƠƯƘƩƞƍŵ
ᲳᲨǭȃȈϋƷᚾᕤƸҗЎƳࣱᏡƕࢽǒǕǔǑƏƴኵǈӳǘƤƯƋǔƷưŴᙌᡯဪӭƷီƳǔᚾᕤǛኵǈӳǘƤƳƍưƘƩƞƍŵ

ᙌᡯᝤ٥Ψ

ᲢEKNᲣ
௾ ங Ⴤ ɦ ᣃ ᝶ ᢼ ᣼ ங ထ ᣼ ஙX [ Z ဪ ‫ע‬ᲫᲪᲨஜᙌԼƷܾ֥Ŵ˄‫ޓ‬ԼሁǛϐМဇӍƸ˂ƷႸႎƴ᠃ဇƠƳƍưƘƩƞƍŵ

ᲬᲩᲬ

loop



ᲶReagentsᲸᲶAmountᲸ

Distilled WaterᲢDWᲣ
2gReaction Mix.ᲢRMᲣ
Primer Mix. DNAᲢPM DNAᲣBst DNA Polymerase

7.0 L
12.5 L
2.5 L1.0 L

Total

23.0 LᲩtest

3ᲣGently tap the tubes for a few timesᲢtappingᲣ, or mix the solution byrepeatedly reversing the tube, or mix by vortex mixer 3 times for 1
second. After mixing well, centrifuge the tubes for a few secondsᲢKeepon iceᲣ.

ᲮᲨOperation procedureᲶMixing of Reagent and SamplesᲸᲢOperate on iceᲣ
Dispense 23.0L of master mix. into each Loopamp Reaction Tube.

Add 2.0L of sample solution and control into the tubes.
      ᲢThe volume of the solution should be 25.0L in total.Უ
Negative controlᲴDistilled WaterᲢDWᲣ
Positive controlᲴPositive control DNAᲢPC DNAᲣMix the solution well by pipetting, or tapping and spin down  ᲢBe careful not to cause airᲧbubbles when mixing.Უ
ᲶAmplificationᲸ
Set the reaction tubes to the reaction block of Loopamp Realtime
Turbidimeter or incubator with hot bonnet, and start the reaction
60᳸65é, 30᳸60 minutes.è5
Control reactionᲴ65é䋬60 minutes
Enzyme inactivationᲢ80éᲦ5 minutes or 95éᲦ2minutesᲣ
ᲢDone by Loopamp Realtime Turbidimeter automatically.Უ
Turbidity measurementȷJudgment
è5ᲴThe reaction condition is dependent upon the characteristics of the primer

for use, therefore examine the optimum condition beforehand.

ᲯᲨDetection
᲻ᲨFor RealᲧtime turbidity detection
RealᲧtime turbidity detection can be carried out with Loopamp Realtime
Turbidimeter. For the detailed operation of the equipment, refer to the
instruction manual of the equipment.
᲼ᲨFor Visual fluorescence detection
Visual fluorescence detection can be achieved by using Loopamp
Fluorescent Detection Reagent4 Უ. For more information, refer to the
package insert for Loopamp Fluorescent Detection Reagent.

ᲫᲩᲬ

3LP2019ᲧE

ᲰᲨAmplification plotsƂCaution for disposalƃ

3LP2019ÜE

ᲫᲨKeep the cap of the used tube completely closed and dispose it, according to
the relevant regulations and instructions, by incineration or after double
bagging it with sealable vinyl bag. To prevent the amplified products fromdispersing, do not conduct autoclave sterilization treatment for2013 ࠰ 10 உμ᩿ોᚖᄂᆮဇᚾᕤ

LAMP ඥƴǑǔ‫ࠢف‬ƴƸᢘЏƳ primer ᚨᚘƕ᣻ᙲƳǫǮƱƳǓǇƢŵᚨᚘƴƋƨƬƯƸŴLAMP ඥ‫ݦ‬ဇƷ primer ᚨᚘૅੲǽȕȈžLAMP ඥȗȩǤȞȸᚨᚘૅੲǽȕȈǦǧ

®

Ǣ Primer ExplorerſǛŴEiken GENOME SITEᲢURLᲵhttp://loopamp.eiken.co.jp/Უ

disposal.
ᲬᲨThe reagent tube is made of polypropylene and the main material for kit
case is paper.
ᲭᲨThe institution disposing the reagent tube and case should bear the
responsibility and abide by the clinical waste disposal regulations, waterpollution prevention law, and any other regulation related.

Storage Expiration

Loopamp DNA Amplification Kit

Ყ20éƂCaution for amplification, detection and judgmentƃ
ᲫᲨLAMP reaction is very sensitive and even the slightest amount of amplified
product tainted into the reaction might cause false result. Therefore,
avoid this type of contamination and carry out the sample and reagent
preparation in different clean benches or separate the area for each
operation. In order to prevent contamination, wear gloves and isolation
gown, etc. if needed.
ᲬᲨWhen this kit used, avoid the contamination of a microbe or nuclease(DNase, RNase).
ᲭᲨNotice that too much mixing by vortex mixer might inactivate Bst DNA
polymerase, and assure that vortexing is conducted 3 times for 1 second.
The prepared master mix. should be used as soon as possible.
ᲮᲨSince bubbles in the solution will interfere the turbidity measurement andcause false judgment, try not to cause any bubble when mixing themaster mix. and the sample solution. If bubbles are present, spin down
to get rid of the bubbles.
ᲯᲨThe cap of the used reaction tubes should not be opened. Contaminationof amplified products on other samples may not only cause false
judgment of the test result but also pollute testing area. In this case, acorrect test result may not be obtained unless pollution is completely
removed.
ᲰᲨDo not carry out electrophoresis of the amplified product.

1 yearǑǓƝМဇƍƨƨƘƜƱƕưƖǇƢŵᲾ᳈᲻ ‫ࠢف‬ᚾᕤ ǭȃȈǇƨŴprimer ӳ঺ƷǰȬȸȉƴƭƍƯƸŴprimer Ʒችᙌࡇƕ᭗ƍǄƲӒࣖᡮࡇƸᡮƘƳǓŴӒࣖᐯ˳ǋ‫ܭܤ‬ƠǇƢŵƠƨƕƬƯŴprimer ƷɟഏǹǯȪȸȋȳǰǛႸႎƱƠஜᙌԼƸᄂᆮဇᚾᕤưƢŵᚮૺӍƸƦƷᙀяǛႸႎƱƠƯƸ̅ဇƠƳƍưƘƩƞƍŵ
ƜƷᛟଢ୿ǛǑƘᛠǜưƔǒ̅ဇƠƯƘƩƞƍŵ

ƂStorageᲦExpirationᲦUnitᲦCode No.ƃProduct NameƯӳ঺Ƣǔ‫ئ‬ӳƸቇତǫȩȠችᙌǰȬȸȉˌɥŴprimer Ǜൿ‫ܭ‬ƢǔᨥӏƼൿ‫ࢸܭ‬ƸŴ‫ݲ‬ƳƘƱǋ FIPᲦBIP ƴƭƍƯƸ HPLC ችᙌǰȬȸȉƴǑǔӳ঺Ǜਖ਼‫ڜ‬ƠǇƢŵᲭᲨᚾᕤƷᛦᙌ૾ඥ

Unit

Code No.

48tests

LMP204

LAMPᲢLoopᲧ mediated Isothermal Amplification ᲣඥƸŴĬ ሁภưᢡˡ‫ࠢف܇‬Ӓࣖ

96tests

LMP205ƕᡶᘍƢǔ 1Უ, 2ᲣᲦĭ 6 ᪸؏ǛᛐᜤƢǔ 4 ᆔ᫏Ʒ primer Ǜ̅ဇƢǔƨǊཎီࣱƕ᭗ƍᲦ 

192tests LMP206

ƂƸơǊƴƃĮ ‫ࠢف‬јྙƕ᭗ƘŴჺ଺᧓ƴ‫ࠢف‬ӧᏡưƋǔᲦį ‫ࠢف‬ငཋ᣽ƕ‫ٶ‬ƘŴቇତ౨ЈƴᢘƠ
Ưƍǔ 3Უ, 4ᲣᲦሁƷཎࣉǛஊƢǔᢡˡ‫ࠢف܇‬ඥưƢŵƂReferencesƃ
1ᲣNotomi T. et al., : Nucleic Acids Research 28, No.12, e 63Ტ2000Უ
2ᲣNagamine K. et al., : Clin. Chem. 47, No. 9, 1742Ყ1743Ტ2001Უ
3ᲣMori Y. et al., : Biochem. Biophys. Res. Commun. 289, No.1, 150Ყ154Ტ2001Უ
4ᲣTomita N. et al., : Nat Protoc. 3, No. 5, 877Ყ882
5ᲣNagamine K. et al., : Molecular and Cellular Probes 16,No.3.223Ყ229
Ტ2002Უ
6ᲣThe guideline for the bioᲧsafety and bioᲧhazardᲢby the Japanese Society
for Bacteriology Უ: Japanese Journal of Bacteriology 54, No.3, 667 Ყ 715
Ტ1999Უ
Licensed under U.S. Patent #5,814,506.

ᲬᲨPrimer Ʒᚨᚘ

1ᲣᲧ20éư̬‫܍‬ƠƯƍƨӲᚾᕤǛܴภưᚐϵƠŴᚐϵࢸƸႺƪƴ൧ɥư̬‫܍‬ƠǇƢŵӲᚾᕤƸŴȁȥȸȖǛ᠉ƘӡƍƯฆӳƢǔᲢǿȃȔȳǰᲣƔŴӍƸ᠃͂ฆԧŴƋǔƍƸ
ȜȫȆȃǯǹȟǭǵȸᲫᅺ᧓gᲭ‫ׅ‬ટਉƠŴǹȔȳȀǦȳƠƯƔǒƝ̅ဇƘƩƞƍŵ
2ᲣȞǹǿȸȟȃǯǹƷᛦᙌᲢ᳖਄ᠲ૞Უ
КᡦဇॖƠƨȞǹǿȸȟȃǯǹᛦᙌဇ๒ᓏȁȥȸȖƴӲᚾᕤǛ࣏ᙲƳȆǹȈૠЎŴɦஜᙌԼƸŴᐯǒᚨᚘƠƨ LAMP ဇȗȩǤȞȸŴǇƨƸК٥ƷȗȩǤȞȸǻȃȈᙌԼƱኵᘙƷлӳᲢ1 ȆǹȈƋƨǓᲣưЎදƠǇƢŵ

ǈӳǘƤƯŴLAMP ඥƴǑǓ೅ႎᢡˡ‫܇‬ᣐЗǛ‫ࠢف‬ȷ౨ЈƢǔƨǊƷǭȃȈưƢŵ

‫ݵ‬ŴȗȩǤȞȸǻȃȈƱኵǈӳǘƤƯ̅ဇƢǔ‫ئ‬ӳƸŴӲȗȩǤȞȸǻȃȈƷ̅ဇᛟଢ୿ƴࢼƬƯƘƩƞƍŵƂϋܾƃ

48 ᴛᴕᴜЎ 96 ᴛᴕᴜЎ 192 ᴛᴕᴜЎᲫᲨ 2 × Reaction Mix.ᲢRMᲣè1

0.6 mL

×1

×2

Ź ǵȳȗȫӒࣖဇᲶᚾᕤᲸ            Ჶဇ᣽ᲸDistilled WaterᲢDWᲣX
L Ტᢘ᣽Უ
2 × Reaction Mix.ᲢRMᲣ
12.5
L
PrimerᲴ FIP
40 pmol
BIP
40 pmol
LFè4
20 pmolLBè4
20 pmolF3
5 pmolB3
5 pmolBst DNA Polymerase
1.0
L

×4

è1

ᲬᲨ Bst DNA PolymeraseᲢBst DNA PolymeraseᲣᲭᲨ Distilled WaterᲢDWᲣè1

60L

×1

×2

×4

1.0 mL

×1

×2

×2

25L

×1

Ყ

Ყ

0.1 mL×1

ᲧᲧᲮᲨ Primer Mix. DNA è2ᲢPM DNAᲣè1
ᲯᲨ Positive Control DNA è2ᲢPC DNAᲣè1
è1ᲴᲢᲣϋƸŴᚾᕤȁȥȸȖƴᚡ᠍ƞǕƯƍǔᘙᅆưƢŵè2Ჴ96 ȆǹȈЎƱ 192 ȆǹȈЎƴƸ Primer Mix. DNA ӏƼ Positive Control DNA ӳ ᚘ

ƸԃǇǕƯƓǓǇƤǜŵ

LAMP ඥƸŴ4 ᆔ᫏Ʒ primer Ʊᦋፗ੭෇ࣱǛਤƭ DNA Polymerase ǛဇƍƯӒࣖǛᘍƏሁภᢡˡ‫ࠢف܇‬ඥưƢŵ4 ᆔ᫏Ʒ primer ƷƏƪŴ2 ᆔ᫏Ʒ inner primer ƸŴƦƷ 3’

ᲫᲨThe kit is designed for research use only.
ᲬᲨWhen handling the sample, always abide by the biohazard countermeasures6Უ.
ᲭᲨWhen ultraviolet lamp is used for the fluorescence visual judgment, do notstare directly at the UV light. Look at it through glass board or protective
goggles.

ჺ጑ƞǕǇƢ 5ᲣŵŹ ஜᙌԼƷǳȳȈȭȸȫӒࣖဇᲶᚾᕤᲸ            Ჶဇ᣽ᲸͨƱ 5’ͨư೅ႎᢡˡ‫܇‬ᣐЗɶƷီƳǔ 2 ᪸؏ǛᛐᜤƢǔ primer ưŴ5’ͨƷᣐЗƸƦƷ 3’ͨƔǒƷ˦ᧈӒࣖưӳ঺ƠƨႻᙀᦋ᪸؏ϋƴǢȋȸȫƢǔǑƏᚨ‫ܭ‬ƠǇƢŵ‫ࠢف‬ӒࣖƸŴƜƷ inner primer ƴǑǓဃ঺ƢǔǹȆȠȫȸȗನᡯƔǒƷᐯࠁ˦ᧈӒࣖƱŴȫȸȗᢿЎƴૼƨƴǢȋȸȫƠƨ inner primer ƔǒƷᦋፗ੭ӳ঺ӒࣖǛጮǓᡉƢƜƱưᡶᘍƠǇƢŵƜǕƴǑǓŴLAMP ඥƸဇƍǔᣞእƕ 1 ᆔ᫏ưƋǔƴǋ᧙ǘǒƣŴሁภƂCaution for handlingƃ

L ᲩȆǹȈ

è4Ჴ࣏ƣƠǋ࣏ᙲƋǓǇƤǜƕŴLoop primer ǛλǕǔƜƱư‫᧓଺ࠢف‬ƕኖ 1 /3 ƴƂย‫ܭ‬Ҿ ྸ ƃƂCaution for handling (hazard prevention)ƃ

23.0

Distilled WaterᲢDWᲣ
2 × Reaction Mix.ᲢRMᲣ
Primer Mix. DNAᲢPM DNAᲣBst DNA Polymerase

7.0
12.5
2.5
1.0

 ӳ ᚘ

23.0 L ᲩȆǹȈ

L
L
L
L

‫ࠢف‬ǛܱྵƠƨ૾ඥưƢŵ

ᲫᲨThis kit should be stored at Ყ20é. To prevent the reagents fromdeterioration, only take out the necessary amount of reagents from thefreezer before use.ᲢNo decline was observed in the kit performance evenafter repeated freezing and thawing for 20 times in the quality control
testing. But, in order to maintain the reagents performance, keep off
unnecessary freezing and thawing.Უ.
ᲬᲨOnly use the specified Loopamp Reaction Tube for turbidity or
fluorescence detection. Other reaction tubes might have different opticaltransparency and can cause misjudgment.
ᲭᲨTake full care when handling reaction tubes, as they are vulnerable toscratches or damages.
ᲮᲨCheck carefully to see if the reaction tubes has any crack or scratch before use.
Crack or scratch on the tube might not only cause false judgment but
also contaminate the equipment. If the tubes are broken inside thereaction block of Loopamp Realtime Turbidimeter, the reaction solution
can spill inside the equipment and cause unrecoverable contamination
malfunctioning.
ᲯᲨDo not expose Loopamp Reaction Tube, master mix. preparation tubes to
UV light. A change in color or degeneration caused by ultraviolet lamp
sometimes results in misjudgment.
ᲰᲨIf the operator does not have the experience or knowledge in the field of
the nucleic acid testing, thereÞ
s a possibility of false judgment. Therefore,make sure that the kit is used under the supervision of the experienced
and knowledgeable technicians.
ᲱᲨEIKEN Chemical Co., Ltd. dose not bear any responsibility for false
judgment or any consequential damage derived from the false judgment
caused by nonᲧcapability problems such as operation error.
ᲲᲨUse the kit before the expiration date, which is labeled on the outer box
ᲢExp. DateᲣ.
ᲳᲨDo not mix different lots.
ᲫᲪᲨDo not use the container of this kit, and accessories to reuse or otherpurposes.

‫ݵ‬ŴӒࣖҾྸƷᛇኬƴƭƍƯƸŴEiken GENOME SITEᲢURLᲵhttp://loopamp.

3ᲣȁȥȸȖǛ᠉ƘӡƍƯฆӳƢǔᲢǿȃȔȳǰᲣƔŴӍƸ᠃͂ฆԧŴƋǔƍƸȜȫȆȃ
ǯǹȟǭǵȸᲫᅺ᧓gᲭ‫ׅ‬ટਉƠŴǹȔȳȀǦȳࢸȞǹǿȸȟȃǯǹƱƠǇƢŵᲢ᳖eiken.co.jp/ᲣǛƝӋༀƘƩƞƍŵ

਄଻ሽᲣƂ̅ဇ૾ඥƃ
ᲫᲨ࣏ᙲƳ֥φȷ֥஬ȷᚾ૰ሁ ᲢஜᙌԼƴƸԃǇǕƯƍǇƤǜƷưКᡦဇॖƠƯƘƩƞƍŵᲣᲮᲨદ˺૾ඥ
źȞǹǿȸȟȃǯǹƷЎදŻᲢ᳖਄ᠲ૞Უ
ӒࣖȁȥȸȖ 1 ஜƋƨǓȞǹǿȸȟȃǯǹ 23.0L ǛЎද1ᲣȞǹǿȸȟȃǯǹᛦᙌဇ๒ᓏȁȥȸȖᲢ 0.5 mL ӍƸ 1.5 mLᲣ2ᲣȞǤǯȭȔȚȃȈ
3Უȕǣȫǿȸ˄ȔȚȃȈȁȃȗᲢDNaseᲦRNase ȕȪȸᲣ
4ᲣLoopamp ӒࣖȁȥȸȖᲢ௿ᄂ҄‫ܖ‬ఇࡸ˟ᅈК٥ԼᲣǵȳȗȫ๋෩ӍƸǳȳȈȭȸȫǛƦǕƧǕ 2.0L ชь         ᲢLAMP Ӓࣖ෩ƱƠƯӳᚘ 25.0LᲣᨏࣱǳȳȈȭȸȫƴƸ Distilled WaterᲢDWᲣǛŴࣱᨗǳȳȈȭȸȫƴƸ Positive Control DNAᲢPC DNAᲣǛ̅ဇ

5ᲣӒࣖȁȥȸȖϬҲဇǢȫȟᙌȩȃǯ
6Უ൧ᲢǯȩȃǷȥǢǤǹᲣӏƼǢǤǹȜȃǯǹ
7Უࣇ᣽ቇତᢒ࣎ೞ
8Უ8 ᡲȞǤǯȭȁȥȸȖဇቇତᢒ࣎ೞ
9ᲣȜȫȆȃǯǹȟǭǵȸ᲻ᲨȪǢȫǿǤȠບࡇ౨ЈȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗᲢLAMP ඥ‫ݦ‬ဇᲣè3᲼ᲨᖩήႸᙻ౨Ј
1ᲣLoopamp ᖩήȷႸᙻ౨ЈᚾᕤᲢ௿ᄂ҄‫ܖ‬ఇࡸ˟ᅈК٥ԼᲣ
2ᲣȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗᲢLAMP ඥ‫ݦ‬ဇᲣè3ŴӍƸǤȳǭȥșȸǿȸᲢภࡇችࡇƕd0.5éˌϋᲴțȃȈȜȳȍȃȈ˄Უ3ᲣȒȸȈȖȭȃǯᲢᣞእ‫෇ڂ‬ဇᲣè3
4Უከ‫ٳ‬ዴༀ‫ݧ‬ᘺፗᲢඬᧈ 240᳸260nmᲦ350᳸370nmᲣè35Უ࠼ࠢƷაᦟӍƸ᧸ᜱ᩿ȔȚȃȆǣȳǰӍƸǿȃȔȳǰƴǑǓฆӳࢸǹȔȳȀǦȳ      Ტ ൢණƕᇌƨƳƍǑƏƴදॖƠǇƢŵᲣź᳆᲻᳇᳊ӒࣖŻȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗӍƸǤȳǭȥșȸǿȸƷӒࣖȖȭȃǯƴǻȃȈƠƯӒࣖǛǹǿȸȈ
60᳸ 65é Ღ30᳸ 60 Ў ᧓ 㶎5
ஜ ᙌ Լ Ʒ ǳ ȳ Ȉ ȭ ȸ ȫ Ӓ ࣖƸ 65éᲦ 60 Ў᧓ᣞእ‫෇ڂ‬Ტ80éᲦ5 Ў᧓ӍƸ 95éᲦ2 Ў᧓ᲣᲢ ȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗưƸᐯѣϼྸƞǕǇƢŵᲣບࡇย‫ ܭ‬ȷ Й ‫ܭ‬è5ᲴᚨᚘƠƨ primer ƴǑƬƯᐱᢘவˑƕီƳǔƷưŴவˑ౨᚛ƕ࣏ᙲƴƳǓǇƢŵ

ᲯᲨ౨Ј᲻ᲨȪǢȫǿǤȠບࡇ౨Ј

è3 ᲴᢘࣖᘺፗŴӒࣖͣഥೞᏡŴከ‫ٳ‬ዴༀ‫ݧ‬ƷவˑƴƭƍƯƸŴEiken GENOME SITEȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗᲢLAMP ඥ‫ݦ‬ဇᲣǛဇƍǔƜƱưŴȪǢȫǿǤȠ౨Јƕ

ᲢURLᲵhttp://loopamp.eiken.co.jp/ᲣǛƝӋༀƘƩƞƍŵ

 ManufacturerưƖǇƢŵદ˺ƷᛇኬƸᘺፗƷช˄૨୿ӏƼӕৢᛟଢ୿ሁǛƝӋༀƘƩƞƍŵ
᲼ᲨᖩήႸᙻ౨ЈК٥Ʒ Loopamp ᖩήȷႸᙻ౨ЈᚾᕤǛဇƍǔƜƱưႸᙻЙ‫ܭ‬ƕӧᏡưƢᲮᲣŵᛇኬƸ

143 Nogi, NogiᲧmachi, ShimotsugaᲧgun, Tochigi 329Ყ0114, JapanᖩήȷႸᙻ౨ЈᚾᕤƷ̅ဇᛟଢ୿ǛƝӋༀƘƩƞƍŵ

Date of Revision: October 1, 2013ᲢVer.5Უ

ᲬᲩᲬ

ᲫᲩᲬ

ᲰᲨAmplification plotsƂCaution for disposalƃ

3LP2019ÜE

ᲫᲨKeep the cap of the used tube completely closed and dispose it, according to
the relevant regulations and instructions, by incineration or after double
bagging it with sealable vinyl bag. To prevent the amplified products fromdispersing, do not conduct autoclave sterilization treatment for2013 ࠰ 10 உμ᩿ોᚖᄂᆮဇᚾᕤ

LAMP ඥƴǑǔ‫ࠢف‬ƴƸᢘЏƳ primer ᚨᚘƕ᣻ᙲƳǫǮƱƳǓǇƢŵᚨᚘƴƋƨƬƯƸŴLAMP ඥ‫ݦ‬ဇƷ primer ᚨᚘૅੲǽȕȈžLAMP ඥȗȩǤȞȸᚨᚘૅੲǽȕȈǦǧ

®

Ǣ Primer ExplorerſǛŴEiken GENOME SITEᲢURLᲵhttp://loopamp.eiken.co.jp/Უ

disposal.
ᲬᲨThe reagent tube is made of polypropylene and the main material for kit
case is paper.
ᲭᲨThe institution disposing the reagent tube and case should bear the
responsibility and abide by the clinical waste disposal regulations, waterpollution prevention law, and any other regulation related.

Storage Expiration

Loopamp DNA Amplification Kit

Ყ20éƂCaution for amplification, detection and judgmentƃ
ᲫᲨLAMP reaction is very sensitive and even the slightest amount of amplified
product tainted into the reaction might cause false result. Therefore,
avoid this type of contamination and carry out the sample and reagent
preparation in different clean benches or separate the area for each
operation. In order to prevent contamination, wear gloves and isolation
gown, etc. if needed.
ᲬᲨWhen this kit used, avoid the contamination of a microbe or nuclease(DNase, RNase).
ᲭᲨNotice that too much mixing by vortex mixer might inactivate Bst DNA
polymerase, and assure that vortexing is conducted 3 times for 1 second.
The prepared master mix. should be used as soon as possible.
ᲮᲨSince bubbles in the solution will interfere the turbidity measurement andcause false judgment, try not to cause any bubble when mixing themaster mix. and the sample solution. If bubbles are present, spin down
to get rid of the bubbles.
ᲯᲨThe cap of the used reaction tubes should not be opened. Contaminationof amplified products on other samples may not only cause false
judgment of the test result but also pollute testing area. In this case, acorrect test result may not be obtained unless pollution is completely
removed.
ᲰᲨDo not carry out electrophoresis of the amplified product.

1 yearǑǓƝМဇƍƨƨƘƜƱƕưƖǇƢŵᲾ᳈᲻ ‫ࠢف‬ᚾᕤ ǭȃȈǇƨŴprimer ӳ঺ƷǰȬȸȉƴƭƍƯƸŴprimer Ʒችᙌࡇƕ᭗ƍǄƲӒࣖᡮࡇƸᡮƘƳǓŴӒࣖᐯ˳ǋ‫ܭܤ‬ƠǇƢŵƠƨƕƬƯŴprimer ƷɟഏǹǯȪȸȋȳǰǛႸႎƱƠஜᙌԼƸᄂᆮဇᚾᕤưƢŵᚮૺӍƸƦƷᙀяǛႸႎƱƠƯƸ̅ဇƠƳƍưƘƩƞƍŵ
ƜƷᛟଢ୿ǛǑƘᛠǜưƔǒ̅ဇƠƯƘƩƞƍŵ

ƂStorageᲦExpirationᲦUnitᲦCode No.ƃProduct NameƯӳ঺Ƣǔ‫ئ‬ӳƸቇତǫȩȠችᙌǰȬȸȉˌɥŴprimer Ǜൿ‫ܭ‬ƢǔᨥӏƼൿ‫ࢸܭ‬ƸŴ‫ݲ‬ƳƘƱǋ FIPᲦBIP ƴƭƍƯƸ HPLC ችᙌǰȬȸȉƴǑǔӳ঺Ǜਖ਼‫ڜ‬ƠǇƢŵᲭᲨᚾᕤƷᛦᙌ૾ඥ

Unit

Code No.

48tests

LMP204

LAMPᲢLoopᲧ mediated Isothermal Amplification ᲣඥƸŴĬ ሁภưᢡˡ‫ࠢف܇‬Ӓࣖ

96tests

LMP205ƕᡶᘍƢǔ 1Უ, 2ᲣᲦĭ 6 ᪸؏ǛᛐᜤƢǔ 4 ᆔ᫏Ʒ primer Ǜ̅ဇƢǔƨǊཎီࣱƕ᭗ƍᲦ 

192tests LMP206

ƂƸơǊƴƃĮ ‫ࠢف‬јྙƕ᭗ƘŴჺ଺᧓ƴ‫ࠢف‬ӧᏡưƋǔᲦį ‫ࠢف‬ငཋ᣽ƕ‫ٶ‬ƘŴቇତ౨ЈƴᢘƠ
Ưƍǔ 3Უ, 4ᲣᲦሁƷཎࣉǛஊƢǔᢡˡ‫ࠢف܇‬ඥưƢŵƂReferencesƃ
1ᲣNotomi T. et al., : Nucleic Acids Research 28, No.12, e 63Ტ2000Უ
2ᲣNagamine K. et al., : Clin. Chem. 47, No. 9, 1742Ყ1743Ტ2001Უ
3ᲣMori Y. et al., : Biochem. Biophys. Res. Commun. 289, No.1, 150Ყ154Ტ2001Უ
4ᲣTomita N. et al., : Nat Protoc. 3, No. 5, 877Ყ882
5ᲣNagamine K. et al., : Molecular and Cellular Probes 16,No.3.223Ყ229
Ტ2002Უ
6ᲣThe guideline for the bioᲧsafety and bioᲧhazardᲢby the Japanese Society
for Bacteriology Უ: Japanese Journal of Bacteriology 54, No.3, 667 Ყ 715
Ტ1999Უ
Licensed under U.S. Patent #5,814,506.

ᲬᲨPrimer Ʒᚨᚘ

1ᲣᲧ20éư̬‫܍‬ƠƯƍƨӲᚾᕤǛܴภưᚐϵƠŴᚐϵࢸƸႺƪƴ൧ɥư̬‫܍‬ƠǇƢŵӲᚾᕤƸŴȁȥȸȖǛ᠉ƘӡƍƯฆӳƢǔᲢǿȃȔȳǰᲣƔŴӍƸ᠃͂ฆԧŴƋǔƍƸ
ȜȫȆȃǯǹȟǭǵȸᲫᅺ᧓gᲭ‫ׅ‬ટਉƠŴǹȔȳȀǦȳƠƯƔǒƝ̅ဇƘƩƞƍŵ
2ᲣȞǹǿȸȟȃǯǹƷᛦᙌᲢ᳖਄ᠲ૞Უ
КᡦဇॖƠƨȞǹǿȸȟȃǯǹᛦᙌဇ๒ᓏȁȥȸȖƴӲᚾᕤǛ࣏ᙲƳȆǹȈૠЎŴɦஜᙌԼƸŴᐯǒᚨᚘƠƨ LAMP ဇȗȩǤȞȸŴǇƨƸК٥ƷȗȩǤȞȸǻȃȈᙌԼƱኵᘙƷлӳᲢ1 ȆǹȈƋƨǓᲣưЎදƠǇƢŵ

ǈӳǘƤƯŴLAMP ඥƴǑǓ೅ႎᢡˡ‫܇‬ᣐЗǛ‫ࠢف‬ȷ౨ЈƢǔƨǊƷǭȃȈưƢŵ

‫ݵ‬ŴȗȩǤȞȸǻȃȈƱኵǈӳǘƤƯ̅ဇƢǔ‫ئ‬ӳƸŴӲȗȩǤȞȸǻȃȈƷ̅ဇᛟଢ୿ƴࢼƬƯƘƩƞƍŵƂϋܾƃ

48 ᴛᴕᴜЎ 96 ᴛᴕᴜЎ 192 ᴛᴕᴜЎᲫᲨ 2 × Reaction Mix.ᲢRMᲣè1

0.6 mL

×1

×2

Ź ǵȳȗȫӒࣖဇᲶᚾᕤᲸ            Ჶဇ᣽ᲸDistilled WaterᲢDWᲣX
L Ტᢘ᣽Უ
2 × Reaction Mix.ᲢRMᲣ
12.5
L
PrimerᲴ FIP
40 pmol
BIP
40 pmol
LFè4
20 pmolLBè4
20 pmolF3
5 pmolB3
5 pmolBst DNA Polymerase
1.0
L

×4

è1

ᲬᲨ Bst DNA PolymeraseᲢBst DNA PolymeraseᲣᲭᲨ Distilled WaterᲢDWᲣè1

60L

×1

×2

×4

1.0 mL

×1

×2

×2

25L

×1

Ყ

Ყ

0.1 mL×1

ᲧᲧᲮᲨ Primer Mix. DNA è2ᲢPM DNAᲣè1
ᲯᲨ Positive Control DNA è2ᲢPC DNAᲣè1
è1ᲴᲢᲣϋƸŴᚾᕤȁȥȸȖƴᚡ᠍ƞǕƯƍǔᘙᅆưƢŵè2Ჴ96 ȆǹȈЎƱ 192 ȆǹȈЎƴƸ Primer Mix. DNA ӏƼ Positive Control DNA ӳ ᚘ

ƸԃǇǕƯƓǓǇƤǜŵ

LAMP ඥƸŴ4 ᆔ᫏Ʒ primer Ʊᦋፗ੭෇ࣱǛਤƭ DNA Polymerase ǛဇƍƯӒࣖǛᘍƏሁภᢡˡ‫ࠢف܇‬ඥưƢŵ4 ᆔ᫏Ʒ primer ƷƏƪŴ2 ᆔ᫏Ʒ inner primer ƸŴƦƷ 3’

ᲫᲨThe kit is designed for research use only.
ᲬᲨWhen handling the sample, always abide by the biohazard countermeasures6Უ.
ᲭᲨWhen ultraviolet lamp is used for the fluorescence visual judgment, do notstare directly at the UV light. Look at it through glass board or protective
goggles.

ჺ጑ƞǕǇƢ 5ᲣŵŹ ஜᙌԼƷǳȳȈȭȸȫӒࣖဇᲶᚾᕤᲸ            Ჶဇ᣽ᲸͨƱ 5’ͨư೅ႎᢡˡ‫܇‬ᣐЗɶƷီƳǔ 2 ᪸؏ǛᛐᜤƢǔ primer ưŴ5’ͨƷᣐЗƸƦƷ 3’ͨƔǒƷ˦ᧈӒࣖưӳ঺ƠƨႻᙀᦋ᪸؏ϋƴǢȋȸȫƢǔǑƏᚨ‫ܭ‬ƠǇƢŵ‫ࠢف‬ӒࣖƸŴƜƷ inner primer ƴǑǓဃ঺ƢǔǹȆȠȫȸȗನᡯƔǒƷᐯࠁ˦ᧈӒࣖƱŴȫȸȗᢿЎƴૼƨƴǢȋȸȫƠƨ inner primer ƔǒƷᦋፗ੭ӳ঺ӒࣖǛጮǓᡉƢƜƱưᡶᘍƠǇƢŵƜǕƴǑǓŴLAMP ඥƸဇƍǔᣞእƕ 1 ᆔ᫏ưƋǔƴǋ᧙ǘǒƣŴሁภƂCaution for handlingƃ

L ᲩȆǹȈ

è4Ჴ࣏ƣƠǋ࣏ᙲƋǓǇƤǜƕŴLoop primer ǛλǕǔƜƱư‫᧓଺ࠢف‬ƕኖ 1 /3 ƴƂย‫ܭ‬Ҿ ྸ ƃƂCaution for handling (hazard prevention)ƃ

23.0

Distilled WaterᲢDWᲣ
2 × Reaction Mix.ᲢRMᲣ
Primer Mix. DNAᲢPM DNAᲣBst DNA Polymerase

7.0
12.5
2.5
1.0

 ӳ ᚘ

23.0 L ᲩȆǹȈ

L
L
L
L

‫ࠢف‬ǛܱྵƠƨ૾ඥưƢŵ

ᲫᲨThis kit should be stored at Ყ20é. To prevent the reagents fromdeterioration, only take out the necessary amount of reagents from thefreezer before use.ᲢNo decline was observed in the kit performance evenafter repeated freezing and thawing for 20 times in the quality control
testing. But, in order to maintain the reagents performance, keep off
unnecessary freezing and thawing.Უ.
ᲬᲨOnly use the specified Loopamp Reaction Tube for turbidity or
fluorescence detection. Other reaction tubes might have different opticaltransparency and can cause misjudgment.
ᲭᲨTake full care when handling reaction tubes, as they are vulnerable toscratches or damages.
ᲮᲨCheck carefully to see if the reaction tubes has any crack or scratch before use.
Crack or scratch on the tube might not only cause false judgment but
also contaminate the equipment. If the tubes are broken inside thereaction block of Loopamp Realtime Turbidimeter, the reaction solution
can spill inside the equipment and cause unrecoverable contamination
malfunctioning.
ᲯᲨDo not expose Loopamp Reaction Tube, master mix. preparation tubes to
UV light. A change in color or degeneration caused by ultraviolet lamp
sometimes results in misjudgment.
ᲰᲨIf the operator does not have the experience or knowledge in the field of
the nucleic acid testing, thereÞ
s a possibility of false judgment. Therefore,make sure that the kit is used under the supervision of the experienced
and knowledgeable technicians.
ᲱᲨEIKEN Chemical Co., Ltd. dose not bear any responsibility for false
judgment or any consequential damage derived from the false judgment
caused by nonᲧcapability problems such as operation error.
ᲲᲨUse the kit before the expiration date, which is labeled on the outer box
ᲢExp. DateᲣ.
ᲳᲨDo not mix different lots.
ᲫᲪᲨDo not use the container of this kit, and accessories to reuse or otherpurposes.

‫ݵ‬ŴӒࣖҾྸƷᛇኬƴƭƍƯƸŴEiken GENOME SITEᲢURLᲵhttp://loopamp.

3ᲣȁȥȸȖǛ᠉ƘӡƍƯฆӳƢǔᲢǿȃȔȳǰᲣƔŴӍƸ᠃͂ฆԧŴƋǔƍƸȜȫȆȃ
ǯǹȟǭǵȸᲫᅺ᧓gᲭ‫ׅ‬ટਉƠŴǹȔȳȀǦȳࢸȞǹǿȸȟȃǯǹƱƠǇƢŵᲢ᳖eiken.co.jp/ᲣǛƝӋༀƘƩƞƍŵ

਄଻ሽᲣƂ̅ဇ૾ඥƃ
ᲫᲨ࣏ᙲƳ֥φȷ֥஬ȷᚾ૰ሁ ᲢஜᙌԼƴƸԃǇǕƯƍǇƤǜƷưКᡦဇॖƠƯƘƩƞƍŵᲣᲮᲨદ˺૾ඥ
źȞǹǿȸȟȃǯǹƷЎදŻᲢ᳖਄ᠲ૞Უ
ӒࣖȁȥȸȖ 1 ஜƋƨǓȞǹǿȸȟȃǯǹ 23.0L ǛЎද1ᲣȞǹǿȸȟȃǯǹᛦᙌဇ๒ᓏȁȥȸȖᲢ 0.5 mL ӍƸ 1.5 mLᲣ2ᲣȞǤǯȭȔȚȃȈ
3Უȕǣȫǿȸ˄ȔȚȃȈȁȃȗᲢDNaseᲦRNase ȕȪȸᲣ
4ᲣLoopamp ӒࣖȁȥȸȖᲢ௿ᄂ҄‫ܖ‬ఇࡸ˟ᅈК٥ԼᲣǵȳȗȫ๋෩ӍƸǳȳȈȭȸȫǛƦǕƧǕ 2.0L ชь         ᲢLAMP Ӓࣖ෩ƱƠƯӳᚘ 25.0LᲣᨏࣱǳȳȈȭȸȫƴƸ Distilled WaterᲢDWᲣǛŴࣱᨗǳȳȈȭȸȫƴƸ Positive Control DNAᲢPC DNAᲣǛ̅ဇ

5ᲣӒࣖȁȥȸȖϬҲဇǢȫȟᙌȩȃǯ
6Უ൧ᲢǯȩȃǷȥǢǤǹᲣӏƼǢǤǹȜȃǯǹ
7Უࣇ᣽ቇତᢒ࣎ೞ
8Უ8 ᡲȞǤǯȭȁȥȸȖဇቇତᢒ࣎ೞ
9ᲣȜȫȆȃǯǹȟǭǵȸ᲻ᲨȪǢȫǿǤȠບࡇ౨ЈȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗᲢLAMP ඥ‫ݦ‬ဇᲣè3᲼ᲨᖩήႸᙻ౨Ј
1ᲣLoopamp ᖩήȷႸᙻ౨ЈᚾᕤᲢ௿ᄂ҄‫ܖ‬ఇࡸ˟ᅈК٥ԼᲣ
2ᲣȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗᲢLAMP ඥ‫ݦ‬ဇᲣè3ŴӍƸǤȳǭȥșȸǿȸᲢภࡇችࡇƕd0.5éˌϋᲴțȃȈȜȳȍȃȈ˄Უ3ᲣȒȸȈȖȭȃǯᲢᣞእ‫෇ڂ‬ဇᲣè3
4Უከ‫ٳ‬ዴༀ‫ݧ‬ᘺፗᲢඬᧈ 240᳸260nmᲦ350᳸370nmᲣè35Უ࠼ࠢƷაᦟӍƸ᧸ᜱ᩿ȔȚȃȆǣȳǰӍƸǿȃȔȳǰƴǑǓฆӳࢸǹȔȳȀǦȳ      Ტ ൢණƕᇌƨƳƍǑƏƴදॖƠǇƢŵᲣź᳆᲻᳇᳊ӒࣖŻȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗӍƸǤȳǭȥșȸǿȸƷӒࣖȖȭȃǯƴǻȃȈƠƯӒࣖǛǹǿȸȈ
60᳸ 65é Ღ30᳸ 60 Ў ᧓ 㶎5
ஜ ᙌ Լ Ʒ ǳ ȳ Ȉ ȭ ȸ ȫ Ӓ ࣖƸ 65éᲦ 60 Ў᧓ᣞእ‫෇ڂ‬Ტ80éᲦ5 Ў᧓ӍƸ 95éᲦ2 Ў᧓ᲣᲢ ȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗưƸᐯѣϼྸƞǕǇƢŵᲣບࡇย‫ ܭ‬ȷ Й ‫ܭ‬è5ᲴᚨᚘƠƨ primer ƴǑƬƯᐱᢘவˑƕီƳǔƷưŴவˑ౨᚛ƕ࣏ᙲƴƳǓǇƢŵ

ᲯᲨ౨Ј᲻ᲨȪǢȫǿǤȠບࡇ౨Ј

è3 ᲴᢘࣖᘺፗŴӒࣖͣഥೞᏡŴከ‫ٳ‬ዴༀ‫ݧ‬ƷவˑƴƭƍƯƸŴEiken GENOME SITEȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗᲢLAMP ඥ‫ݦ‬ဇᲣǛဇƍǔƜƱưŴȪǢȫǿǤȠ౨Јƕ

ᲢURLᲵhttp://loopamp.eiken.co.jp/ᲣǛƝӋༀƘƩƞƍŵ

 ManufacturerưƖǇƢŵદ˺ƷᛇኬƸᘺፗƷช˄૨୿ӏƼӕৢᛟଢ୿ሁǛƝӋༀƘƩƞƍŵ
᲼ᲨᖩήႸᙻ౨ЈК٥Ʒ Loopamp ᖩήȷႸᙻ౨ЈᚾᕤǛဇƍǔƜƱưႸᙻЙ‫ܭ‬ƕӧᏡưƢᲮᲣŵᛇኬƸ

143 Nogi, NogiᲧmachi, ShimotsugaᲧgun, Tochigi 329Ყ0114, JapanᖩήȷႸᙻ౨ЈᚾᕤƷ̅ဇᛟଢ୿ǛƝӋༀƘƩƞƍŵ

Date of Revision: October 1, 2013ᲢVer.5Უ

ᲬᲩᲬ

ᲫᲩᲬ

ᲰᲨ‫୺ࠢف‬ዴȑǿȸȳƂࡑూɥƷදॖƃFor research use onlyᲬᲨPrimer design
 Appropriate primer design is essential for amplification by LAMP. For
the primer design, exclusive primer designing software, PrimerExplorer isavailable from Eiken GENOME SITE (URL;http://loopamp.eiken.co.jp/e/).
When use the highly purified primers, a rapid gene amplification canbe performed and stable reproducibility of the amplification can beobtained. The first screening for appropriate LAMP primers might notnecessarily require highly purified primers. However, after the primers
are determined, it is recommended to use purified FIP and BIP through
HPLC or better purification.

ᲫᲨӒࣖࢸƷȁȥȸȖƸᔟǛ᧏ƚƣƴŴ໲ҲϼྸӍƸ݅᧍ưƖǔȓȋȸȫƷᘥǛʚ᣻ƴ଀®

ƠᢘЏƴϼྸƠƯƘƩƞƍŵ‫ࠢف‬ငཋƷ᫠૝᧸ഥƷƨǊŴᑄ᫈ߩ㓙ߦࠝ࡯࠻ࠢ࡟࡯

ࡉಣℂߪⴕࠊߥ޿ߢߊߛߐ޿‫ޕ‬ᲬᲨᚾᕤȁȥȸȖƸ᳊᳊ŴǭȃȈǱȸǹƸኡǛɼƳ஬ឋƱƠƯƍǇƢŵ
ᲭᲨ̅ဇࢸƷஜԼǍܾ֥ӏƼ֥φ᫏ƸŴࡑూཋƷϼྸӏƼฌੈƴ᧙ƢǔඥࢷǍ൦ឋ൲ບ᧸DNA Amplification KitഥඥሁƴࢼƍŴӲ଀ᚨƷᝧ˓ƴƓƍƯϼྸƠƯƘƩƞƍŵ

This kit is for research use only. Do not use fordiagnosis or its assistance for clinical purpose.
Read this instruction carefully before use.

Ƃᝪᔺ૾ඥȷஊј஖᧓ȷѼᘺҥˮȷᙌԼǳȸȉƃᙌ Լ Ӹᝪᔺ૾ඥ

Loopamp DNA ‫ࠢف‬ᚾᕤǭȃȈ

Ყ20 é

ஊј஖᧓

1 ࠰᧓

Ჶย‫ܭ‬ƴƋƨƬƯƷදॖᲸ
ᲫᲨLAMP ӒࣖƸ᩼ࠝƴᥘ૓ƳӒࣖưƋǓŴ೅ႎᢡˡ‫܇‬Ǎ‫ࠢف‬ငཋƕಊࣇ᣽ưǋฆλƢǔƱᛚƬƨኽௐǛǋƨǒƢҾ‫׆‬ƱƳǔƓƦǕƕƋǓǇƢŵƜƷǑƏƳǳȳǿȟȍȸǷȧ

Ѽᘺҥˮ

ᙌԼǳȸȉ

48 ȆǹȈЎ

LMP20496 ȆǹȈЎ

LMP205

192 ȆǹȈЎ

LMP206

Ƃɼᙲ૨ྂƃ

ȳǛ‫ׅ‬ᢤƢǔƨǊƴŴ౨˳੔ӕƸஜᙌԼǛ̅ဇƢǔᢿ‫ދ‬ƱƸКƷᢿ‫ދ‬ưܱ଀ƢǔƔŴ1ᲣNotomi T. et al., : Nucleic Acids Research 28, No.12, e 63Ტ2000ᲣƋǔƍƸ౨௹ғ؏ǛЎлƠƯКƷǨȪǢưᘍƬƯƘƩƞƍŵ࣏ᙲƴࣖơƯŴǯȪȸȳ

2ᲣNagamine K. et al., : Clin. Chem. 47, No.9, 1742Ყ1743Ტ2001Უ

șȳȁƷ̅ဇӍƸ৖ᘥǍǢǤǽȬȸǷȧȳǬǦȳƷბဇሁưǳȳǿȟȍȸǷȧȳǛ

3ᲣMori Y. et al., : Biochem. Biophys. Res. Commun. 289, No.1, 150Ყ154Ტ2001Უ
4ᲣTomita N.et al., : Nat Protoc. 3, No.5, 877Ყ882᧸ƙਜ਼ፗǛƱƬƯƘƩƞƍŵ

5ᲣNagamine K. et al., : Molecular and Cellular Probes 16, No.3, 223Ყ229Ტ2002ᲣᲬᲨஜᙌԼǛӕǓৢƏᨥƴƸŴࣇဃཋǍఋᣠЎᚐᣞእᲢDNaseᲦRNaseᲣƷǳȳǿȟȍȸ

6Უଐஜኬᓏ‫˟ܖ‬ȐǤǪǻǤȕȆǣȸ‫ۀ‬Ճ˟Ჴଐஜኬᓏ‫ܖ‬ᩃᛏ, 54, No.3, 667Ყ715Ტ1999Უ

ǷȧȳǛᢤƚƯƘƩƞƍŵ
ᲭᲨBst DNA Polymerase Ƹ‫෇ڂ‬ƢǔƓƦǕƕƋǓǇƢƷưŴȞǹǿȸȟȃǯǹᛦᙌƷᨥƴƸນƠƘટਉƠƳƍưƘƩƞƍŵȜȫȆȃǯǹȟǭǵȸưટਉǛᘍƏ‫ئ‬ӳƸŴ1 ᅺᲭᲨReagents preparationƂCharacteristicsƃ
LAMP Ტ Loop Ყ mediated Isothermal Amplification Უ method is a gene
amplification method capturing the following characteristics : Ĭ The
amplification reaction proceeds under isothermal condition1 Უ, 2 Უ.,ĭ It has
extremely high specificity because of the use of 4 primers recognizing 6
distinct regions on the target.,Į It has high amplification efficiency andenables amplification within a shorter time., and į it produces tremendous
amount of amplified products which makes simple detection possible3Უ, 4Უ.
  This kit is for amplifying and detecting target gene sequence by the
LAMP method in combination with personally designed primers for LAMP, orprimer sets sold separately.

Ƃ Contents of the kitƃ     1Შ 2 × Reaction Mix.ᲢRMᲣè1௿ᄂ҄‫ܖ‬ఇࡸ˟ᅈ ƓܲಮႻᛩᆸӝ3Შ Distilled WaterᲢDWᲣè1ȕȪȸȀǤȤȫ㩷 㩷 㩷 0120Ყ308Ყ421

ƞƍŵŹ Sample reaction
48 tests 96 tests 192 tests

0.6 mLg1   g2    g4

4Შ Primer Mix. DNA è 2ᲢPM DNAᲣè1ᲮᲨȞǹǿȸȟȃǯǹƱǵȳȗȫ๋෩ǛฆԧࢸŴӒࣖ෩ƴൢණƕസƬƯƍǔƱບࡇย‫ܭ‬Ʒ

5Შ Positive Control DNA è 2ᲢPC DNAᲣè1

60L

g1   g2    g4

1.0 mLg1   g2    g2

25Lg1  Ყ   Ყ

0.1 mLg1  Ყ   ᲧᲶReagentsᲸ

Distilled WaterᲢDWᲣ
2gReaction MixᲢRMᲣ      PrimerᲴ FIP
BIP
LFè4
LBè4
F3
B3
Bst DNA Polymerase2Შ Bst DNA PolymeraseᲢBst DNA PolymeraseᲣè1ƂբƍӳǘƤέƃ

᧓g3 ‫ׅ‬Ǜӈ‫ܣ‬ƠƯƘƩƞƍŵ‫ݵ‬ŴᛦᙌƠƨȞǹǿȸȟȃǯǹƸƢƙƴ̅ဇƠƯƘƩ  

1Უ Take out the reagents stored at Ყ 20 é , and thaw them at room
temperature. Once the reagents are thawed, keep them on ice. After
thawed, tap the reagent tubes gently -- mixing (tapping) -- or invert thetube, or by vortex mixer at about 3 times for 1 second. After mixing well,centrifuge the tubes for a few seconds.
2Უ Preparation of master mix.ᲢOperate on iceᲣ.
The following amount of the component is required for one reaction.
When used in combination with Loopamp Primer Set, follow the
preparation instructions of each primer set.
ᲶAmountᲸ   
X
12.5
40
40
20
20
5
5
1.0

Total

L (ad q.s.)
L
pmol
pmol
pmol
pmol
pmol
pmol
L

23.0

LᲩtestૅᨦƱƳǓᛚЙ‫ܭ‬ƷҾ‫׆‬ƱƳǓǇƢƷưŴൢණƕဃơƳƍǑƏදॖƠƯƘƩƞƍŵൢè1ᲴThe notation on each reagent tube is shown inᲢණƕസƬƯƍǔ‫ئ‬ӳƴƸŴǹȔȳȀǦȳƠƯൢණǛӕǓᨊƍƯƘƩƞƍŵ

Უ.

è4ᲴLoop primers are not necessarily required. However, the use ofprimers shortens the amplification time by about one-third5Უ.

è2ᲴPrimer Mix. DNA and Positive Control DNA are not included in the 96ᲯᲨӒࣖࢸƷȁȥȸȖƷᔟƸൿƠƯ᧏ƚƳƍưƘƩƞƍŵཎƴӒࣖࢸƷȁȥȸȖǛᘺፗƔ

tests and 192 tests package.

ǒӕǓЈƢƱƖƴȁȥȸȖƷᔟƕ᧏ƔƳƍǑƏŴ॰᣻ƴӕǓЈƠƯƘƩƞƍŵ‫ࠢف‬ငŹ Control reactionƂPrincipleƃ

ཋƴǑǔǳȳǿȟȍȸǷȧȳƸᛚЙ‫ܭ‬ƷҾ‫׆‬ƱƳǔƹƔǓưƳƘŴย‫ܭ‬࿢‫ؾ‬ƦƷǋƷ

LAMP method is an isothermal nucleic acid amplification method using
4 kinds of primers and DNA polymerase with strand displacement activity.
Among 4 kinds of primers, two of them are inner primers, whose 3’ end and
5’ end are respectively designed to be complementary to two different
regions of the target sequence. When the primerᲧlinked strand is replicated,
resulting 3Þend structure to selfᲧanneals to the complementary region in selfᲧ
structure, it forms the loop structure at the end.
This stemᲧloop structure allow the 3Þ end to initiate selfᲧelongation andanother inner primer to anneal to its loop region to synthesize new DNA
strand with strandᲧdisplacement manner. Through the repetition of theseprocesses, the amplification proceeds, and this enables amplification tocontinue under isothermal condition with only one kind of enzyme.
For further details of the LAMP method, refer to Eiken GENOME SITEᲢURLᲵhttp://loopamp.eiken.co.jp/e/Უ.

Ǜ൲௨ƠŴ൲௨Ǜ‫ܦ‬μƴᨊӊƠƳƍᨂǓŴˌࢸദƠƍኽௐƕࢽǒǕƳƘƳǔӧᏡࣱƕƋǓǇƢŵᲰᲨᩓൢපѣሁưƷ‫ࠢف‬ငཋƷӕৢƍƸᢤƚƯƘƩƞƍŵ

ƂӕৢƍɥᲢүᨖ᧸ഥᲣƷදॖƃᲫᲨஜᙌԼƸŴ˳‫ٳ‬ᚮૺဇҔᕤԼưƸƋǓǇƤǜŵ
ᲬᲨ౨˳Ƹज़௨ƷүᨖƕƋǔǋƷƱƠƯදॖƠƯӕǓৢƍŴ࣏ᙲƳȐǤǪȏǶȸȉ‫ݣ‬ሊǛܱ଀ƠƯƘƩƞƍ 6ᲣŵᲭᲨᖩήႸᙻЙ‫଺ܭ‬ƴከ‫ٳ‬ዴༀ‫ݧ‬ᘺፗǛ̅ဇƢǔ‫ئ‬ӳŴȩȳȗǑǓ્‫ݧ‬ƞǕǔከ‫ٳ‬ዴᲢൈᓏዴᲣƸஊܹƳƷưŴከ‫ٳ‬ዴǛႺ੗ႸƴλǕǔƜƱƸᢤƚƯƘƩƞƍŵǇƨໜ໊ɶƷȩȳȗǛᙸǔ࣏ᙲƕƋǔƱƖƸŴ࣏ƣǬȩǹெǛᡫƢƔŴ࠼ࠢƷაᦟӍƸ᧸ᜱ᩿ǛƔƚƯЙ‫ܭ‬ƠƯƘƩƞƍŵ

ƂHow to useƃƂ̅ဇɥƷදॖƃ

ᲫᲨMaterials required but not provideᲫᲨஜᙌԼƸ࣏ƣᲧ20éư̬‫܍‬ƠƯƘƩƞƍŵᚾᕤƷэ҄Ǜ᧸ഥƢǔƨǊƴŴ̅ဇ଺Ƹ࣏

1ᲣSterilized tubes for master mix. preparationᲢ0.5 mL or 1.5 mLᲣ
2ᲣMicropipettes
3ᲣPipette tips with filterᲢDNase, RNase freeᲣ4ᲣLoopamp Reaction TubeᲢsold separatelyᲣ5ᲣAluminum rack for cooling tubes
6ᲣIceᲢcrushed iceᲣand ice box
7ᲣCentrifuge for microtube
8ᲣCentrifuge for 8Ყstrip tubes
9ᲣVortex mixerᙲƳᚾᕤƩƚǛረƔǒӕǓЈƠƯƝ̅ဇƘƩƞƍŵ
ᲢϵኽᗡᚐǛ 20 ‫ׅ‬ጮǓᡉƠƨኽௐưƸŴᚾᕤƷэ҄ƸǄƱǜƲᛐǊǒǕƯƓǓǇƤǜƕŴ໯ဇƳϵኽᗡᚐƸԼឋዜਤƷƨǊƴᢤƚƯƘƩƞƍŵᲣᲬᲨӒࣖȁȥȸȖƸ࣏ƣ‫ݦ‬ဇƷ Loopamp ӒࣖȁȥȸȖǛƝ̅ဇƘƩƞƍŵਦ‫ٳˌܭ‬ƷӒࣖȁȥȸȖǛ̅ဇƠƨ‫ئ‬ӳŴήᡢᢅࣱƷᢌƍƴǑǓᛚЙ‫ܭ‬ǛਔƘӧᏡࣱƕƋǓǇƢŵ
ᲭᲨӒࣖȁȥȸȖƸᄊ੷ƠǍƢƍƷưŴӕৢƍƴƸදॖƠƯƘƩƞƍŵ
ᲮᲨӒࣖȁȥȸȖƸဇƍǔЭƴǭǺȷȒȓሁƕ໯ƍƜƱǛႸᙻưᄩᛐƠƯƘƩƞƍŵӒࣖ

᲻ᲨFor RealᲧtime turbidity detection
Loopamp Realtime Turbidimeter è3ȁȥȸȖƴǭǺȷȒȓሁƕƋǔƱദƠƘย‫ܭ‬ưƖƳƍƹƔǓƔŴȁȥȸȖƷᄊ੷ƴǑǓᘺፗǛ൲௨ƢǔӧᏡࣱƕƋǓǇƢŵȪǢȫǿǤȠບࡇย‫ܭ‬ᘺፗӍƸǤȳǭȥșȸǿȸưŴஜᙌԼƷ̅ဇƴƋƨƬƯƸᢡˡ‫܇‬౨௹ƷჷᜤŴኺ᬴ǛஊƠƨ২ᘐᎍƷਦ‫ݰ‬ƷǋƱ

᲼ᲨFor Visual fluorescence detection
1ᲣLoopamp Fluorescent Detection ReagentᲢsold separatelyᲣ2ᲣLoopamp Realtime Turbidimeter è3, or incubator with hot bonnet
Ტtemperature accuracy withind0.5éᲣ3ᲣHeat blockᲢfor enzyme inactivationᲣè3
4ᲣUV lampᲢwavelength at 240᳸260nm or 350᳸370nmᲣè3
5ᲣProtective goggle or glass boardư౨௹Ǜܱ଀ƠƯƘƩƞƍŵ

è3ᲴFor the information about applicable instrument, reaction terminationƷӒࣖȖȭȃǯϋưȁȥȸȖƕᄊ੷Ơƨ‫ئ‬ӳƸŴӒࣖ෩ƕᘺፗஜ˳ǁ๨ЈƠŴᨊӊɧᏡƳ൲௨Ǎ૏ᨦƷҾ‫׆‬ƱƳǓǇƢŵ
ᲯᲨӒࣖȁȥȸȖŴȞǹǿȸȟȃǯǹᛦᙌဇ๒ᓏȁȥȸȖƴƸከ‫ٳ‬ዴༀ‫ݧ‬ƠƳƍưƘƩƞ
ƍŵከ‫ٳ‬ዴༀ‫ݧ‬ƴǑǔ‫٭‬ᑥȷ‫٭‬ឋሁưᛚƬƨኽௐǛǋƨǒƢ‫ئ‬ӳƕƋǓǇƢŵ
ᲰᲨᢡˡ‫܇‬౨௹ƷჷᜤǍኺ᬴ǛǋƨƳƍ‫ئ‬ӳŴ౨௹ኽௐƷЙ‫ܭ‬ǛᛚǔүᨖࣱƕƋǓǇƢƷ

function and condition for UV irradiation, refer to Eiken GENOME SITEᲢURL;http://loopamp.eiken.co.jp/e/Უ.

ᲱᲨஜᙌԼƷࣱᏡƴဌஹƠƳƍʙဌᲢદ˺૾ඥǛᛚƬƨ‫ئ‬ӳሁᲣƴǑǔᛚƬƨЙ‫ܭ‬ŴǇƨƦƷЙ‫ܭ‬ƴဌஹƠƯႆဃƠƨʙ᪮ƴ‫ݣ‬ƠƯŴࡴᅈƸɟЏƷᝧ˓Ǜ᝟ƍǇƤǜŵ
ᲲᲨஜᙌԼƸ̅ဇ஖ᨂϋƴ̅ဇƠƯƘƩƞƍŵ
ᲳᲨǭȃȈϋƷᚾᕤƸҗЎƳࣱᏡƕࢽǒǕǔǑƏƴኵǈӳǘƤƯƋǔƷưŴᙌᡯဪӭƷီƳǔᚾᕤǛኵǈӳǘƤƳƍưƘƩƞƍŵ

ᙌᡯᝤ٥Ψ

ᲢEKNᲣ
௾ ங Ⴤ ɦ ᣃ ᝶ ᢼ ᣼ ங ထ ᣼ ஙX [ Z ဪ ‫ע‬ᲫᲪᲨஜᙌԼƷܾ֥Ŵ˄‫ޓ‬ԼሁǛϐМဇӍƸ˂ƷႸႎƴ᠃ဇƠƳƍưƘƩƞƍŵ

ᲬᲩᲬ

loop



ᲶReagentsᲸᲶAmountᲸ

Distilled WaterᲢDWᲣ
2gReaction Mix.ᲢRMᲣ
Primer Mix. DNAᲢPM DNAᲣBst DNA Polymerase

7.0 L
12.5 L
2.5 L1.0 L

Total

23.0 LᲩtest

3ᲣGently tap the tubes for a few timesᲢtappingᲣ, or mix the solution byrepeatedly reversing the tube, or mix by vortex mixer 3 times for 1
second. After mixing well, centrifuge the tubes for a few secondsᲢKeepon iceᲣ.

ᲮᲨOperation procedureᲶMixing of Reagent and SamplesᲸᲢOperate on iceᲣ
Dispense 23.0L of master mix. into each Loopamp Reaction Tube.

Add 2.0L of sample solution and control into the tubes.
      ᲢThe volume of the solution should be 25.0L in total.Უ
Negative controlᲴDistilled WaterᲢDWᲣ
Positive controlᲴPositive control DNAᲢPC DNAᲣMix the solution well by pipetting, or tapping and spin down  ᲢBe careful not to cause airᲧbubbles when mixing.Უ
ᲶAmplificationᲸ
Set the reaction tubes to the reaction block of Loopamp Realtime
Turbidimeter or incubator with hot bonnet, and start the reaction
60᳸65é, 30᳸60 minutes.è5
Control reactionᲴ65é䋬60 minutes
Enzyme inactivationᲢ80éᲦ5 minutes or 95éᲦ2minutesᲣ
ᲢDone by Loopamp Realtime Turbidimeter automatically.Უ
Turbidity measurementȷJudgment
è5ᲴThe reaction condition is dependent upon the characteristics of the primer

for use, therefore examine the optimum condition beforehand.

ᲯᲨDetection
᲻ᲨFor RealᲧtime turbidity detection
RealᲧtime turbidity detection can be carried out with Loopamp Realtime
Turbidimeter. For the detailed operation of the equipment, refer to the
instruction manual of the equipment.
᲼ᲨFor Visual fluorescence detection
Visual fluorescence detection can be achieved by using Loopamp
Fluorescent Detection Reagent4 Უ. For more information, refer to the
package insert for Loopamp Fluorescent Detection Reagent.

ᲫᲩᲬ

3LP2019ᲧE