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last updated: 2025-12-14 RT after BA fixed inconsistencies
file_name: SOP-104-A_1 Test Training - Run Form.md
file_date: 2022-08-26
title: FloodLAMP QMS SOP-104-A_1 Test Training - Run Form
category: guides
subcategory: qms-sops
tags: 
source_file_type: gdoc
xfile_type: docx
gfile_url: https://docs.google.com/document/d/1Nk586b_UbYKr6iE7Yrhe7NHN1tl4qSttFPYquUHJQJQ
xfile_github_download_url: https://raw.githubusercontent.com/FocusOnFoundationsNonprofit/floodlamp-archive/main/guides/qms-sops/SOP-104-A_1%20Test%20Training%20-%20Run%20Form.docx
pdf_gdrive_url: https://drive.google.com/file/d/1JmcisdqvDVhxCmCFEHRi3YacFW9xYaBB
pdf_github_url: https://github.com/FocusOnFoundationsNonprofit/floodlamp-archive/blob/main/guides/qms-sops/SOP-104-A_1%20Test%20Training%20-%20Run%20Form.pdf
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license: CC BY 4.0 - https://creativecommons.org/licenses/by/4.0/
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words: 1021
notes: 
summary_short: The FloodLAMP QMS SOP-104-A_1 is an amplification training run form that walks trainees through contamination-control practices, 1XISS preparation, sample inactivation, reaction mix preparation, and the 25-minute LAMP amplification workflow. It includes a checklist-style procedure with specific handling rules (e.g., glove changes, control addition order, timing, and imaging settings) to prevent post-amplification contamination. The form provides write-in fields for batch/heater/reagent/control IDs and lookup tables for tracking sample positions in strip tubes or plates.


CONTENT

FloodLAMP Biotechnologies
**Document Number:** SOP-104-A_1
**Effective Date:** 08/26/2022
**Version:** 1

***INTERNAL TITLE:*** Amplification Training - Run Form
## PREP
 - Pen & sharpie labeled with green tape
 - Separation of Post-Amp area
 - Heaters on (Heat indicator lit!)
 - Safety Procedures:
   - lab coat
   - gloves
   - face mask
   - face shield or goggles
 - Alcohol wipe sample tubes

## PREP USER WRITE IN
Log Link: SOP-104-B_1 {SITE}
Location:
Name:
Date:
Time:

## CONTAMINATION INFRACTIONS
-   PPE lapse/removal
-   Use green labeled pen/marker with bare hands and not putting it to be cleaned afterwards
-   Use unmarked pen/marker with clean gloves (without cleaning gloves after or changing gloves)
-   Bare hands on anything improper, except in desk area
-   Reuse pipette tip improperly
-   Touch pipette tip to anything and then not discard it
-   Touch anything dirty then go back to work (face, mask, uncleaned phone, uncleaned laptop)

## NOTE
It is recommended that 1XISS and Reaction Mix be prepared before the assay run (instructions below).

## DOCUMENTATION
-   Check off each item after completing it
-   Write legibly

## GLOSSARY OF TERMS
**100XIS**………………………….. 100-fold concentrated Inactivation Solution
**1XISS**……………………………. 1X Inactivation Saline Solution
**PGS**………………………………. Primer Guanidine Solution
**CLAMP MM**…………………. Colorimetric Loop-Mediated Isothermal Amplification Master Mix
**RM**………………………………… Reaction Mix (CLAMP MM and PGS)
**TPC**………………………………. Twist Positive Control

## 1XISS PREP
 - Record volumes of saline & 100XIS in 1XISS Prep form prior to pipetting
 - Combine saline and 100XIS in a 50mL falcon tube or 30mL Chub tube, depending on volume
 - Vortex 30 seconds
 - Label tube with 1XISS ID (e.g., "1X_MMDD-1") and record the same ID into the 1XISS Prep Form and the table on this form

## 1XISS PREP USER WRITE IN
Sample Batch ID (from App):
Inactiv Heater:
1X Inactiv Saline Soln ID:
Dispense: Pipette or Manual Disp or Electric Disp

## INACTIVATION
-   Add 1mL of 1XISS to each sample tube
-   Verify that tubes are properly sealed afterward (i.e., not cross-threaded)
-   Vortex 10 sec (if individual tubes) or 30 sec (if tubes-in-rack)
-   Heat 8 min in Water Bath (set to 99°C)
-   Cool 10 min
-   Put in refrigerator before 30 minute mark

## REACTION MIX PREP
-   Complete the entry in the RM Prep Form prior to pipetting
-   Calc volumes correctly for PGS and CLAMP based on number of reactions
-   Remove CLAMP MM and PGS from freezer to thaw, but do not allow them to warm
-   Briefly spin down reagent tubes (removes reagent from the cap and tube walls)
-   Pipette CLAMP MM and PGS into a 1.5mL tube using calculated volumes
-   Promptly return PGS and CLAMP source tubes to freezer
-   Vortex and spin down the RM tube
-   Aliquot 23uL of RM into each reaction tube (plate or strips-of-8)
    -   Pipetting at the bottom of tubes

    -   No blowout while dispensing reaction mix (to prevent bubbles in bottom of well)
-   Visually check the volumes in each reaction tube (that they appear to be even)
-   If using a plate, cover filled tubes with rack lid to prevent dust
-   At this stage, RM reaction tubes can be capped and placed in the freezer for storage if they are not to be used fresh.

## AMPLIFICATION REACTION
- Remove PCR cold block from freezer ~10 min before adding tubes
- Prep RM plate/strips (thawed, not warm)
- Set up tips to align with reactions
- Use a lookup rack to keep track of sample order
  - Tip layout aligns with arrangement of sample tubes in lookup rack & aligns with layout of reaction plate/strips
- Add negative control first (2uL 1XISS)
- Add 2uL of each inactivated sample to reaction tubes, visually checking the volume in the tip each time. Pipette up & down 5X, blow out in liquid, tip touch.
- With each add, verify the correct position (tip vs sample tube vs reaction tube)
- Transfer plate/strip to PCR rack (not cold block - rxns don't go to amp cold)
- Retrieve TPC from freezer, touching only with one hand (consider this hand dirty)
- Add 2uL TPC to pos. ctrl, return TPC to freezer (IMMEDIATELY change glove)
- Cap reaction tubes / seal plate
- Change gloves again after loading tubes/plate onto amp heater
- Set main timer for 25 min
- Set secondary timer for 24 min
- Put sample tubes in fridge
- Remove rxns from amp promptly, wait 1 minute before imaging
- Take photo in lightbox
- For photo apply crop and 'Vivid' filter (or increase saturation 70% for android)
- Log amp run sheet with QR code
- On app: Intake, Process and Result tubes

## AMPLIFICATION REACTION USER WRITE IN
Amp Heater:  
Strip8 Tubes or Plates  
Reaction Mix: Frozen or Fresh  
Reaction Mix ID:  
Num Reactions (including controls):  
Pos Ctrl ID:  
Neg Ctrl ID (today's 1XISS):  
Initial Inconclusives?:  
Notes:

|         |       |       |       |       |       |       |
|---------|-------|-------|-------|-------|-------|-------|
|         | **1** | **2** | **3** | **4** | **5** | **6** |
| **A 1** |       |       |       |       |       |       |
| **B 2** |       |       |       |       |       |       |
| **C 3** |       |       |       |       |       |       |
| **D 4** |       |       |       |       |       |       |
| **E 5** |       |       |       |       |       |       |
| **F 6** |       |       |       |       |       |       |
| **G 7** |       |       |       |       |       |       |
| **H 8** |       |       |       |       |       |       |
||

|         |       |       |       |        |        |        |
|---------|-------|-------|-------|--------|--------|--------|
|         | **7** | **8** | **9** | **10** | **11** | **12** |
| **A 1** |       |       |       |        |        |        |
| **B 2** |       |       |       |        |        |        |
| **C 3** |       |       |       |        |        |        |
| **D 4** |       |       |       |        |        |        |
| **E 5** |       |       |       |        |        |        |
| **F 6** |       |       |       |        |        |        |
| **G 7** |       |       |       |        |        |        |
| **H 8** |       |       |       |        |        |        |
||