Synthesis · Development of an electrochemiluminescence aptasensor combining covalent-triazine framework emitter with exonuclease III-driven DNA walker for sensitive CEA detection

7 structured synthesis routes

Completeness describes how fully the route could be reconstructed from the main article and supporting information.

Complete recipeSource: SI

Route 1: Other

SI text · S1.5 Pre-treatment of the bare AE

Metal precursorsgold-disk electrode

Solventswater; 0.5 M H2SO4 electrolyte

Additives0.05 um aluminium oxide polishing powder

Atmosphereambient

Substrate orientationgold-disk electrode

Oxidant / reductantelectrochemical cycling from -0.2 V to +1.6 V

Work-upMechanical polishing, water sonication, electrochemical polishing for 25 cycles at 100 mV s-1.

Scalability contextStandard electrode pre-treatment; one-electrode analytical workflow.

Show 3 structured reagent records

Role	Reagent	Amount / concentration	Source
Other	gold-disk electrode	Not specified	SI text · S1.5 Pre-treatment of the bare AE
Other	aluminium oxide	0.05 um particle size	SI text · S1.5 Pre-treatment of the bare AE
Electrolyte	H2SO4	0.5 M	SI text · S1.5 Pre-treatment of the bare AE

Partial recipeSource: Both

Route 2: Drop Cast

2-3 · 2.2 Construction of the CTF-based ECL sensor

Linker precursorsCTF powder

SolventsCTF suspension solvent not specified in text layer

Atmosphereair drying

Substrate orientationgold electrode (AE)

Work-upWashed with Milli-Q water three times to remove loosely stacked material.

Scalability contextDrop-cast analytical electrode; no film-thickness or scale-up data.

Show 2 structured reagent records

Role	Reagent	Amount / concentration	Source
Other	CTF suspension	10 uL · 0.15, 0.25, 0.5, 0.6, 0.8, 1.0, and 1.5 mg mL-1 tested; 0.8 mg mL-1 optimal	2-3 · 2.2 Construction of the CTF-based ECL sensor
Other	gold electrode (AE)	Not specified	2-3 · 2.2 Construction of the CTF-based ECL sensor

Complete recipeSource: Both

Route 3: Solvothermal

2 · 2.1 The preparation of CTF

Linker precursorsTTB (19.7 mg, 0.05 mmol); TTA (17.7 mg, 0.05 mmol)

Solvents1,4-dioxane, mesitylene, and 3 M glacial acetic acid, v/v/v = 5:5:1, total volume 1.1 mL

Additives3 M glacial acetic acid

AtmosphereSchlenk tube; degassed through three freeze-pump-thaw cycles

Temperature120

Time72

Work-upYellow precipitate collected by centrifugation and thoroughly rinsed with ethanol and acetone.

ActivationDried under vacuum overnight.

Scalability contextSmall-batch Schlenk-tube synthesis; no scale-up data reported.

Show 5 structured reagent records

Role	Reagent	Amount / concentration	Source
Linker	4,4',4''-(1,3,5-triazine-2,4,6-triyl)tribenzaldehyde (TTB)	19.7 mg, 0.05 mmol	2 · 2.1 The preparation of CTF
Linker	4,4',4''-(1,3,5-triazine-2,4,6-triyl)trianiline (TTA)	17.7 mg, 0.05 mmol	2 · 2.1 The preparation of CTF
Solvent	1,4-dioxane	part of 1.1 mL solvent mixture, v/v/v = 5:5:1	2 · 2.1 The preparation of CTF
Solvent	mesitylene	part of 1.1 mL solvent mixture, v/v/v = 5:5:1	2 · 2.1 The preparation of CTF
Acid	glacial acetic acid	part of 1.1 mL solvent mixture, v/v/v = 5:5:1 · 3 M	2 · 2.1 The preparation of CTF

Partial recipeSource: SI

Route 4: Hydrothermal

SI text · S1.4 Preparation of CuxMn3-x(HITP)2

Metal precursorsCoCl2.6H2O (6.6 mg, as printed in SI, inconsistent with Cu product); MnCl2.6H2O (6.6 mg)

Linker precursorsHITP (10 mg)

Solventswater (10 mL)

Additivesaqueous ammonia (0.6 mL)

Atmospherenot specified

Temperature120

Time24

Oxidant / reductantaqueous ammonia added after dissolution; no separate oxidant/reductant specified

Work-upPrecipitate collected and washed with acetone.

ActivationDried in vacuum oven; drying temperature/time not specified for pristine powder.

Scalability contextSmall solution batch; no scalability data reported.

Show 5 structured reagent records

Role	Reagent	Amount / concentration	Source
Metal Source	CoCl2.6H2O (as printed; likely conflicts with Cu product)	6.6 mg	SI text · S1.4 Preparation of CuxMn3-x(HITP)2
Metal Source	MnCl2.6H2O	6.6 mg	SI text · S1.4 Preparation of CuxMn3-x(HITP)2
Linker	2,3,6,7,10,11-hexaiminotriphenylene (HITP)	10 mg	SI text · S1.4 Preparation of CuxMn3-x(HITP)2
Solvent	water	10 mL	SI text · S1.4 Preparation of CuxMn3-x(HITP)2
Base	aqueous ammonia	0.6 mL	SI text · S1.4 Preparation of CuxMn3-x(HITP)2

Complete recipeSource: Both

Route 5: Other

3 · 2.2 Construction of the CTF-based ECL sensor · Scheme 1

Metal precursorsHp-labelled CuxMn3-x(HITP)2 probe

Linker precursorsMCH/ssDNA/CTF/AE, CEA aptamer, cDNA, Hp DNA/rHp

SolventsPBS; dsDNA/CEA incubation mixture

AdditivesExo III; CEA analyte; K2S2O8 only for measurement, not assembly

Atmospherenot specified

Temperature37; 80 for enzyme inactivation

Time1 h aptamer/cDNA dsDNA formation; 2 h CEA incubation; 1.5 h Exo III incubation; 10 min at 80 C

Substrate orientationMCH/ssDNA/CTF on gold electrode

Work-upThe ssDNA/CTF/AE was soaked in the Exo-III-digested Hp/cDNA-functional MOF solution.

ActivationHeated to 80 C for 10 min to inactivate Exo III.

Scalability contextAnalytical biosensor assembly; no large-area/device scalability reported.

Show 6 structured reagent records

Role	Reagent	Amount / concentration	Source
Other	CEA-target aptamer	10 uL · 200 nM	3 · 2.2 Construction of the CTF-based ECL sensor · Scheme 1
Other	cDNA	10 uL · 200 nM	3 · 2.2 Construction of the CTF-based ECL sensor · Scheme 1
Other	CEA solution	10 uL · varied	3 · 2.2 Construction of the CTF-based ECL sensor · Scheme 1
Other	Hp-labeled CuxMn3-x(HITP)2	30 uL · 100 ng mL-1 probe stock reported in SI	3 · 2.2 Construction of the CTF-based ECL sensor · Scheme 1
Other	Exo III	100 U	3 · 2.2 Construction of the CTF-based ECL sensor · Scheme 1
Other	MCH/ssDNA/CTF/AE	one electrode	3 · 2.2 Construction of the CTF-based ECL sensor · Scheme 1

Complete recipeSource: SI

Route 6: Other

SI text · S1.7 Immobilization of the Hp on the CuxMn3-x(HITP)2

Metal precursorsCuxMn3-x(HITP)2

Linker precursorsHp DNA strand

SolventsHp strand solution; PBS for redispersion

Atmospherenot specified

Temperature35 for vacuum drying

Time6 h soaking; 24 h vacuum drying

Substrate orientationpowder suspension

Work-upCollected by centrifugation and washing.

ActivationVacuum dried at 35 C for 24 h, then redispersed in PBS.

Scalability contextProbe prepared at milligram scale for analytical sensing.

Show 3 structured reagent records

Role	Reagent	Amount / concentration	Source
Other	CuxMn3-x(HITP)2	5 mg	SI text · S1.7 Immobilization of the Hp on the CuxMn3-x(HITP)2
Other	Hp strand solution	0.5 mL · 10 uM	SI text · S1.7 Immobilization of the Hp on the CuxMn3-x(HITP)2
Solvent	PBS	10 mL for redispersion	SI text · S1.7 Immobilization of the Hp on the CuxMn3-x(HITP)2

Partial recipeSource: Main

Route 7: Other

2-3 · 2.2 Construction of the CTF-based ECL sensor

Linker precursorsssDNA on CTF; MCH blocking molecule

SolventsPBS (0.1 M, pH 7.4); MCH solution

Additives6-mercapto-1-hexanol (MCH)

Atmosphereair drying

Temperature4 for storage

Time1 h ssDNA incubation; MCH soaking time not specified

Substrate orientationCTF/AE

Work-upRinsed with PBS to remove unanchored strand; air-dried after MCH blocking.

ActivationStored at 4 C for subsequent use.

Scalability contextAnalytical electrode modification.

Show 3 structured reagent records

Role	Reagent	Amount / concentration	Source
Other	ssDNA strand solution	10 uM	2-3 · 2.2 Construction of the CTF-based ECL sensor
Other	6-mercapto-1-hexanol (MCH)	1 uM	2-3 · 2.2 Construction of the CTF-based ECL sensor
Solvent	PBS	0.1 M, pH 7.4	2-3 · 2.2 Construction of the CTF-based ECL sensor